ACTA PROTOZOOLOGICA VOLUME 19

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ACTA PROTOZOOLOGICA
VOLUME 19
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EXPERIMENTAL
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PROTOZOOLOGICA
International Journal of Protozoology
Editors
Stanisław DRYL and Stanisław L. KAZUBSKI
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Munaging
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ACTA
Vol.. 19 (INo. 1)
PHOTOZOOLOGICA
WARSZAWA, 30 III 1980
pp. 1-14
Department of Biology, Wayne State University, Detroit, Michigan 48202, USA
Gregory
A. A N T I P A 1
A Temporal Analysis of Cell Cycle and Morphogenetic
2 3
Events in Tetrahymena
pyriformis
'
Received
15 April 1978, accepted
30 August 1979
Synopsis. The relative duration of oral morphogenesis and DNA
synthesis has been investigated by an approach which combines the
methodology of the chemos+at with a demographic analysis. Details of
the method are presented. Cell generation times were varied under
constant conditions of temperature and nutrient limitation while cells
w e r e maintained in balanced growth. Data collected at generation times
of 3 to 14 h indicate that the morphogenetic sequence is unaltered and
most closely approaches a time-dependent assembly process. DNA synthesis appears to be a cycle-dependent phenomenon and requires an
interval which is proportional to the total cycle time. Thus, controls
for oral morphogenesis and DNA synthesis do not appear to be tightly
linked. These results are dependent upon the assumptions of the method
and hypotheses tested.
In recent years oral morphogenesis or stomatogenesis of the ciliated
protozoa has been examined with increased interest in an effort to
establish phylogenetic relationships among related organisms (see e.g.,
C o r l i s s 1968, P e c k 1974, S m a l l 1967). From such studies emerges the notion of a constitutive set of ordered morphological events.
That is, one observes a progression or sequence of specific events (stages) for a given organism, and stage B, not only logically, but always
follows stage A, etc. There is a clear impression that ciliate stomato1
Present address: Department of Biology, San Francisco State University,
San Francisco, California 94132.
2
This project was supported by United States Public Health Services Grant
S 0 8 RR07051 awarded by the Biomedical Research Development Grant Program,
Division of Research Resources, National Institutes of Health.
* An earlier report of material contained in this paper was presented at
the 5th International Congress of Protozoology in New York on June 28, 1977.
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G. A N T I P A
genesis is a carefully prescribed developmental sequence and as such
may be a "programmed" process. The possibility that this morphogenetic program is an intimate part of the cell cycle is worthy of our
consideration.
Although many studies have detailed the morphological stages of
stomatogenesis, there have been but a few which have directed attention
to the time-course of events during this dynamic process (F r a n k e 1
1960, 1964, H o l z 1960, W i l l i a m s 1964). Further, while it has been
well established that ciliates will have different generation times in
different growth media ( W i l l e 1972) or at different temperatures
(T h o r m a r 1959, G i e s e and M c C a w 1963), no detailed analysis
of the temporal events of stomatogenesis has yet been carried out under
conditions of varying generation time 4. One hint of what might be expected comes from F r a n k e l ' s 1960 analysis of Glaucoma chattoni.
Fig. 1. Circular chronogram diagramatically represents time-dependent and cycledependent hypotheses. Perimeter distance represents real time-in-cycle. Note that
time-in-cycle is identical for both conditions only at the double circle
* After the preliminary parts of this report were completed, w e were informed of the work of S u h r - J e s s e n et al. which has subsequently been published (1977). The Discussion of the present paper has been revised to consider
this important contribution.
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T E M P O R A L A N A L Y S I S OF C E L L U L A R
EVENTS
3
Here he provides some data on the time of stomatogenesis as found in
three different strains of G. chattoni which differed in their rates of
growth. He concludes that "...the a b s o l u t e duration of stomatogenesis
is relatively constant and species-specific...."
While Frankel provides us with the suggestion that stomatogenesis
may be a time and species dependent phenomenon, another alternative
is that stomatogenesis is intimately linked with the cell cycle, and as
cells move to longer generation times, these events "stretch" out in
a proportional manner; the cycle-dependent hypothesis. In their simplest forms, the time-dependent and cycle-dependent hypotheses are
diagrammed in Fig. 1. Although M i t e h i s o n (1974) discusses a variety of ways that cell life cycle events may be linked, the time- or
cycle-dependent hypotheses will suffice for our present considerations.
We have developed a novel approach to this problem (but see footnote 4) which employs the technology of the chemostat coupled with
a demographic analysis of the data. In this way it was possible to collect data on one strain of Tetrahymena pyriformis grown at one temperature but with a variety of specific limiters to growth and generation times. We have ascertained absolute values for the duration of s o matogenic stages under these conditions. From this analysis it was possible to get an idea about the plasticity and limits of the controlling
mechanism(s) of stomatogenesis. In addition, this method is also applicable to the analysis of other cellular events, and we present preliminary data here for a temporal analysis of DNA synthesis during the cell
cycle.
Materials
and
Methods
Steady-state cultures of axenic Tetrahymena
pyriformis
W were grown at
generation times (GT) of three to twenty-three hours, a 7 2/3-fold range of
GT's. The three hour GT was accomplished in batch culture, and repetitive samples were taken to assure the steady-state conditions of exponential growth.
Other GT's were achieved at 28°C in a conventional N o v i c k and S z i l a r d
(1950) chemostat where nutrient limitation was by carbon, leucine or oxygen.
Medium A of H o 1 z et al. (1959) was used without glucose supplementation to
achieve carbon limitation, with l/10th leucine concentration for leucine limitation,
and it was bubbled with 2% oxygen-balance nitrogen for oxygen limitation. In
each case population kinetics were carefully monitored by cell counts made with
a Coulter Counter (Model B) fitted with a 140 nm aperture.
Stages of stomatogenesis were quantitatively scored from C h a t t o n - L w o f f
silver impregnations ( C o r l i s s 1953) of cells taken from each experimental
condition. All slides with poorly impregnated specimens were discarded regardless of the source of their imperfection. In every case, the orientation of specimens scored was rigorously controlled so both the proter buccal area as well
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4
G. A N T I P A
the region of potential or developing opisthe anlagen were clearly visible. All
organisms not meeting this criterion were summarily discounted and are not
reflected in our quantitative data. The seven stages of stomatogenesis {0 and
1-VI) were after F r a n k e l (1964).
In order to estimate the time of DNA synthesis (see below) average DNA
contents were determined on replicate samples of approximately 5 X 10® cells
by a modification of the S c h m i d t and T h a n n h a u s e r (1945) method. Samples
were taken from a steady-state, carbon-limited chemostat (as above), and data
are presented in relative but arbitrary units. A detailed description of the analytical method as well as absolute estimates for T. pyriformis
will appear elsewhere (A n t i p a et al. in preparation).
Analysis of Real Time-in-Cycle
As is the case in all asynchronous and exponentially expanding populations
which do not exhibit cell death, the analysis of events for real time-in-cycle
requires: (1) knowledge of the timing of the event, and (2) adjustment of raw
data for the phenomenon of recruitment from behind. Since one dividing cell
will give rise to two new cells at t0, a growing population will contain a larger
percentage of new cells than old cells. The characteristic and constant population skew which results has been described previously (e.g., P o w e l l 1956,
B o s t o c k 1970, B a r f o r d and H a l l 1976). Since the chemostat provides the
opportunity to investigate a culture which remains in steady-state and asynchronous exponential growth ( K u b i t s c h e k 1970), the population profile within
a chemostat is also described by this characteristic population. It seems,
Fig. 2. Age distribution of idealized population after J a m e s (1974). Nomograph
below allows for conversion of percent observed to percent of cycle time
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TEMPORAL A N A L Y S I S OF CELLULAR
EVENTS
5
appropriate, then, to review the population profile of an asynchronous, exponentially growing population (Fig. 2). Mere we provide the breakdown of an idealized
population, the formula which describes the population based on the logistic
growth curve, and an iteration of this formula which provides a nomograph for
direct conversion of appropriate raw data to time-of-cycle values.
Both unlimited steady-state growth in a batch culture and limited steadystate growth in a chemostat guarantee asynchronous growth, a necessary assumption
for correction of data by the method described above. Further, W i l l i a m s
(1964), P r e s c o t t (1957) and others have demonstrated that individual generation times of T. pyriformis
show but small variation from the mean population
GT. Variability in GT's may be as small as 7%, and w e have no reason to
believe that variability under our chemostat conditions exceeds this estimate.
One consequence of the "tightness" of GT's is that the theoretical population
profile (Fig. 2) closely approximates what one finds in the experimental situation.
We have assumed perfect agreement to the idealized populations in our computations which appear below; the small but inevitable variation from this in our
experiments does not materially influence our results.
Methods of temporal analysis by the digestion of data from asynchronous
cultures provides certain advantages over analysis of either single cell data or
data derived from either synchronous or synchronized cultures. The main advantages are: (1) the ability to simultaneously acquire large numbers of organisms in
all stages of morphogenesis, (2) the opportunity to examine organisms taken
from a homogeneous environment, and (3) especially pertinent to this study,
the compatibility of the analytical method to the experimental methodology of
the chemostat. Not unexpectedly, such a method also has distinct disadvantages.
The principal disadvantage lies in the fact that one must know the position
of the temporal events being investigated within the total cycle time. In the case
of stomatogenesis, since the terminal event (end of stage VI) is also coincident
with the termination of the division cycle (by definition, F r a n k e l 1964) and
since the stomatogenic sequence is a continuous morphogenetic series, this particular problem is alleviated. One simply works backward in the analysis of the
complete series.
When knowledge of the precise positioning of an event is not available, it
is still possible to test data against theoretical considerations, and this is how
we have dealt with our analysis of DNA synthesis. Again, knowledge of the
characteristic population skew is essential, but in this case it provides a predictable effect on the average parameters of the culture, and this provides the
opportunity for testing data against prediction. J a m e s (1974) has presented us
with an important conceptual extention of the population skew in presenting
his concept of the "unit cell." The unit cell refers to the integrated volume of
an irregular cubic with dimensions of time-of-cycle, population number (both
as in Fig. 2) and amount of substance per cell (e.g., DNA/cell). The integrated
volume of this cubic represents the a v e r a g e per cell measurement such as
those we have made on DNA. As a consequence, it is possible to make a series
of assumptions, generate values based on those assumptions, and then compare
experimentally derived data against these predicted values. We present one such
argument in the discussion.
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6
G. A N T I P A
R e s u l t s
Stomatogenesis
Data on the duration of stomatogenesis were collected from eight
different experimental situations. Haw data were analyzed according
to the methods described above, and a summary of these observations
appears in Table 1. Perusal of these data immediately indicates that
there is no major alteration in the course of stomatogenesis under these
experimental conditions. In all cases, all six stages of oral development
have been observed. Further, the sequence of stages does not appear
to be greatly influenced by either the limiter to growth or the GT, as
stages II, III and VI tended to be of short duration in each case while
stages I, IV and V tended to be of longer duration. Stage III was generally of the shortest duration, and stages I, IV and V were longer and,
on the average, of approximately similar duration. In general, the total
time course of stomatogenesis required a smaller portion of the total
cycle time as cells moved to longer generation times. Further, the morphological stages of stomatogenesis required approximately the same
time regardless of the overall cell generation time.
DNA Content
The average per cell DNA content for Tetrahymena grown under
carbon-limitation at generation times of 5.5 to 23 h is given in Table 2.
Our measurements were unable to detect a significant change in DNA
content over this four-fold range of GT's. The significance of this result
is discussed below.
Discussion
We have been able to demonstrate that T. pyrijormis can be maintained for extended periods in continuous exponential and steady-state
growth at generation times of 5 to 23 h. Successful chemostat culture
has been achieved with leucine, oxygen and carbon-limitation, and
under each of these conditions, T. pyrijormis performed admirably by
maintaining a steady and constant titer while meeting conditions of the
imposed generation time. None of the difficulties frequently associated
with the chemostat culture of other organisms ( P i r t 1975) such as cell
death, wall growth or incomplete division proved to be of consequence in
these experiments.
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TEMPORAL A N A L Y S I S OF CELLULAR
7
EVENTS
Table 1
Sample
1
Limiter
UL
GT
3.0
0
n
3
4
LEU
LEU
o2
5.0
5.2
6.4
1000
1000
300
6
7
8
o2
LEU
LEU
C
6.4
9.0
9.5
10.0
300
1000
1000
1000
VI
19.8
50
5.8
10.4
104
12.5
22.5
98
12.5
22.5
73
10.0
18.0
58.2
104.7
c
687
60.7
182.1
78
9.2
27.6
38
4.3
12.9
25
3.1
9.3
74
9.6
28.8
37
4.4
13.2
61
8.7
26.1
39.3
117.9
A
B
C
754
68.3
213.1
75
9.0
28.1
42
5.6
17.5
27
3.1
9.6
47
6.5
20.3
31
4.3
13.4
24
3.2
10.0
31.7
98.9
A
B
209
61.8
237.3
26
10.2
39.2
17
6.8
26.1
4
1.4
5.4
19
8.5
32.6
18
8.0
30.7
7
3.3
12.7
38.2
146.7
c
217
65.0
249.5
16
6.2
23.7
10
3.8
14.5
12
5.0
19.1
16
7.0
26.8
17
7.6
29.8
12
5.4
20.6
35.0
134.5
A
B
C
837
78.2
422.3
39
4.7
25.4
21
3.0
16.2
21
2.9
15.7
30
4.1
22.1
34
4.6
24.8
18
2.5
13.5
21.8
117.7
A
B
C
869
82.3
469.1
23
2.9
16.5
15
2.3
13.1
17
2.1
12.0
25
3.3
18.8
26
3.6
20.5
25
3.5
20.0
17.7
100.9
A
B
C
816
75.8
454.8
43
5.2
31.2
24
3.2
19.2
21
3.0
18.0
30
3.9
23.4
52
7.0
42.0
14
1.9
11.4
24.2
145.2
A
B
c
5
V
Duration
of oral
development
64
6.4
11.5
1000 A
B
c
2
Stomatogenic Stages
I
II
III
IV
A
B
500
41.8
75.3
111
11.0
—
—
—
—
—
—
—
—
Rows A, B and C are respectively: raw data, duration as percent of cycle time, and
duration in minutes. Other abbreviations axe as follows: GT — generation time in hours,
n — number of specimens scored, UL — "unlimited" growth, LEU — leucine, Oa — oxygen,
C — carbon.
The ability to manipulate cell generation times in a specific medium
and at a specified temperature has allowed us to perturb the "normal"
time course of the cell division cycle and examine the relationship between temporal events of stomatogenesis and DNA synthesis. Under
conditions of batch culture growth, for every round of DNA synthesis
there is one sequence of oral morphogenesis; these events remain tightly
coupled. It has been known, however, that conditions for heat shock
synchrony of Tetrahymena result in the uncoupling of these processes,
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8
G. A N T I P A
and DNA synthesis persists while stomatogenesis is blocked (Z e u t h e n
1974, L o w y and L e i c k 1969). What we have found in this study
is a temporal dissociation of events of DNA synthesis and oral morphogenesis under conditions of balanced growth.
Table 2
Sample
Limiter
GT
Relative
DNA
Content
1
2
3
4
5
C
C
5.5
8.5
10.0
14.0
23.0
0.421
0.416
0.411
0.432
0.403
c
c
c
Abbreviations as in Table 1
Stomatogenesis
The results of our experiments on stomatogenesis indicate that this
morphological s e q u e n c e of events takes place in approximately
the same length of time regardless of the cell generation time. This
suggests that stomatogenesis in T. pyriformis, or at least the associated
morphological events, is a fixed sequence that resembles a time-dependent assembly process. Our data are summarized in a linear chronogram
(Fig. 3 A) which indicates the variability in our data but clearly suggests the time-dependent nature of this process. An examination of
Table 1 and Fig. 3 A indicates that the duration of stomatogenesis under
conditions of either carbon or oxygen limitation tends to be longer than
under leucine limitation. Carbon or oxygen limitation may both represent forms of energy limitation while leucine represents amino acid
or nitrogen limitation. These differences may result in the discrepancies
observed, and further experimentation should validate this secondary
phenomenon associated with the timing of stomatogenesis.
S u h r - J e s s e n et al. (1977) have recently performed a remarkably similar experiment. They examined T. pyriformis GL in a chemostat culture with an unspecified limiter. They clearly establish that the
cells are undergoing balanced growth with a single limiter at generation times of 4.9 to 17.2 h. In their study they also conclude that the
course of stomatogenesis is a time-dependent phenomenon, in this case
of approximately 91 m duration at 28°C. We have summarized their
data in Fig. 3 B. It is evident that our data do not precisely agree with
theirs, but this may be explained by either the different strains and
media used or by some differences in the method of scoring. We would
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Fig. 3. Linear chronograms which indicate the onset of stomatogenesis (symbol) at the position observed within the total
cycle time. Numerals at left of each figure indicate the GT in hours. A — data presented here. B — after S u h r - J e s s e n et al. (1977)
T E M P O R A L A N A L Y S I S O F C E L L U L A R E V E N T S 11
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10
G. A N T I P A
like to point out, however, that the time course they have established
most closely fits our samples of either leucine limitation or "unlimited"
growth. This suggests to us that the limitation to growth experienced
by T. pyrijormis in a proteose peptone-based chemostat or in an "unlimited" batch culture may represent either amino acid or nitrogen
limiting conditions.
DNA Synthesis
Since H o w a r d and P e l c (1951) first used autoradiography to
demonstrate that there was a discrete stage during which DNA was
synthesized, innumerable studies on all conceivable types of eukaryotic
organisms have supported this observation, and their subsequent algorithm, Gi — S — G2 — M, has become a precept. Further, comparative
investigations have suggested that under conditions of variable GT, the
most flexible period is G! while S and G2 intervals tend to be constant
in their duration ( M i t c h i s o n 1971). We wish to emphasize that this
"general principle" is based on comparative observations of different
cells or tissues, different strains, cells at different temperatures, or
strains growing in different media. Our results can be interpreted to
deviate from this basic theme.
We have examined T. pyrijormis W grown at the s a m e temperature, in the s a m e medium and under the s a m e conditions of nutrient limitation. We have found no appreciable difference in the a v e r a g e DNA content per cell. With suitable assumptions and application
of the J a m e s (1974) unit cell concept, we can estimate the relative
position of the DNA synthetic interval. This method does not require
the use of isotope incorporation and as such is independent of problems
associated with that method ( M o s e r 1967, C o 11 r e 11 1977).
If we use the customary assumption that DNA synthesis proceeds
at a uniform rate, and if we assume DNA synthesis to occupy one hour
in the middle of a three hour Tetrahymena cell cycle, then we will be
filling the conditions for which direct measurements have been made
( M c D o n a l d 1958, 1962). But, what happens when the generation
time varies under constant conditions of temperature and limiter to
growth? Four generalized alternatives will be considered: (1) S and G2
intervals remain of fixed duration, increase in cycle time appears in
G1( (2) Gi and S intervals remain fixed and the increase in cycle time
appears as an extention of G2, (3) S remains in the center of the cycle
and of fixed duration, Gi and G2 intervals increase proportionally, and
(4) S interval remains in the center of the cycle but increases along
with G t and G2 as the cycle time increases (cycle-dependent hypothesis,
see Fig. 1). Examples for each of these four conditions are depicted in
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TEMPORAL A N A L Y S I S OF CELLULAR
11
EVENTS
Fig. 4. Figure 5 demonstrates the effect the age distribution and assumptions have on the mean DNA content for asynchronous cultures under
each of these four conditions. It will be seen that while each condition
predicts identical values for the mean DNA content at the three hour
GT, conditions 1 and 2 deviate in a predictable and systematic curvilinear fashion for other GT's while conditions 3 and 4 predict a constant
DNA content across generation times. By this method it is not possible
to distinguish between conditions 3 and 4.
Our data conform to the results predicted by conditions 3 and 4
(Fig. 6). Within the context of our experiment, it is possible to distinguish between these two cases with a simple labeling experiment. From
Fig. 4 it will be seen that as cells grow at longer GT's under condition 3,
fewer cells will be involved in S at any given time, while under condition 4, the same proportion is engaged in DNA synthesis at all GT's.
As a consequence, it is possible to resolve between two situations by
©
®
Fig. 4. Diagraraatic representation of four possible conditions (1-4) for the
interval (blocks) at 3, 6, 12 and 24 h GT's. See text for further detail
»z
T
1
LLI
f-
1
1
1
1
1
1
\
oc_> 1.5
1
1
S
1
(3) and (4)
<
z
Q
I
I
I
—1
6
12
GT (hours)
Fig. 5. Average DNA content as a function of generation time for each of the
four conditions described in Fig. 4 and text
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12
G. A N T I P A
0.6
<
Z
JL
04
O
02
0
5
10
15
20
25
30
35
GT (hours)
Fig. 6. Relative but arbitrary DNA content observed in a carbon-limited chemostat (solid circles) in this study and as derived from S u h r - J e s s e n et al.
(1977) (open circles). Line without data points represents condition 1 (above)
and is included for reference
looking at the relative numbers of cells incorporating 3 H-thymidine during a short pulse. If cells are growing according to condition 3, fewer
cells will incorporate 3 H-thymidine at longer GT's than at shorter GT's,
but the intensity of incorporation will be the same in each case. If S
follows the pattern described by condition 4, the same proportion of
cells will incorporate 3 H-thymidine at all GT's, but grain counts per cell
will become less at longer generation times.
Although we have not yet performed this experiment, S u h r - J e s s e n et al. (1977) have investigated the timing of S by conventional
methods in their chemostat study. They conclude that the S-period increased slightly while Gi and G2 periods both increased with the same
proportion at increased generation times. We have analyzed their data
by the method of J a m e s (1974) and find amazing agreement between
their data and ours (see Fig. 6). Our preliminary conclusion and present working hypothesis, based on the data of S u h r - J e s s e n et al.
(1977) and ours, is that the S-period of T. pyrijormis is controlled in
a manner which is proportional to the cell cycle time, hence it is a cycle
dependent phenomenon.
Conclusions
We have demonstrated that the chemostat as analyzed by demographic methods, and especially the unit cell concept of J a m e s (1974),
can be an effective tool for the analysis of temporal events which occur
during the cell division cycle. While we have discussed here the use
of these methods in the analysis of DNA synthesis and stomatogenesis,
it should be emphasized that the methodology is appropriate for the
analysis of any component which doubles during the cell cycle. Since
the chemostat is ideally suited for the examination of balanced growth
during which cellular components double, it is hoped that this approach
will be put to use in other studies of cellular growth.
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TEMPORAL ANALYSIS OF CELLULAR
EVENTS
13
We observe that stomatogenesis most closely approaches a timedependent phenomenon (Fig. 1), and in this conclusion we are in agreement with F r a n k e l (1960) and S u h r - J e s s e n et al. (1977). DNA
synthesis is found to approximate a cycle-dependent phenomenon (Fig.
1), and here we are supported by S u h r - J e s s e n et al. (1977). It
becomes evident, then, that controls for stomatogenesis and DNA synthesis in Tetrahymena are not tightly coupled. It remains the task of
future endeavors to elaborate on these circumstances.
ACKNOWLEDGEMENTS
It is my pleasure to acknowledge the technical assistance provided by B. W.
Swanson, M. Zarski and P. Szczepański during various phases of this project.
The opportunity to examine the Suhr-Jessen et al. (1977) manuscript prior to its
publication is especially appreciated; for this I am indebted to Dr. Leif Rasmussen.
RÉSUMÉ
Le temps relative de la morphogénèse de l'appareil oral et de la synthèse
de l'ADN a été étudié par un procédé qui combine la méthodologie de chemostate
avec l'analyse démographique. Les détails de cette méthode sont décrits. Le temps
de duré des générations des cellules est variables dans les conditions thermiques
et nutritives stables, quand les cellules sont maintenues dans la croissance équilibrée. Les données concernant les temps de génération entre 3 et 14 h montrent que la séquence morphogénétique reste intacte qu'elle approche à un processus dépendant du temps. La synthèse de l'ADN paraît être phénomène dépendant de la phase du cycle et elle nécessite un délai proportionnel au temps total
du cycle. Donc, les mécanismes de contrôle de la morphogénèse de l'appareil
oral et de la synthèse de l'ADN probablement n e sont pas étraitement liés entre
eux.
REFERENCES
B a r f o r d J. P. and H a l l R. J. 1976: Estimation of the length of the cell cycle
phases from asynchronous cultures of Saccharomyces
cerevisiae.
Expl. Cell
Res., 102, 276-284.
B o s t o c k C. J. 1970: DNA synthesis in the fission yeast
Schizosaccharomyces
Plombe. Expl. Cell Res., 60, 16-26.
C o r l i s s J. O. 1953: Silver impregnation of ciliated protozoa by the ChattonLwoff technic. Stain Tech., 28, 97-100.
C o r l i s s J. O. 1968: The value of ontogenetic data in reconstructing protozoan p h y logenies. Trans. Am. Micros. Soc., 87, 1-20.
C o t t r e l l S. F. 1977: Mitochondrial DNA synthesis in synchronous cultures of
the yeast, Saccharomyces
cerevisiae. J. Cell Biol., 75, 15 a.
F r a n k e l J. 1960: Morphogenesis in Glaucoma chattoni. J. Protozool., 7, 362-376
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14
G. A N T I P A
F r a n k e l J. 1964: Cortical morphogenesis and synchronization in
Tetrahymena
pyriformis GL. Expl. Cell Res., 35, 349-360.
G i e s e A. C. and M c C a w B. 1963: Regeneration rate of Blepharisma with special reference to the effect of temperature. J. Protozool., 10, 173-182.
H o l z G. G. Jr. 1960: Structural and functional changes in a generation in Tetrahymena. Biol. Bull., 118, 84^95.
H o l z G. G. Jr., E r w i n J. A. and D a v i s R. J. 1959: Some physiological characteristics of the mating types and varieties of Tetrahymena
pyriformis.
J. Protozool., 6, 149-156.
82
H o w a r d A. and P e l c S. R. 1951: Nuclear incorporation of P as demonstrated
by autoradiographs. Expl. Cell Res., 2, 178-187.
J a m e s T. W. 1974: A cell cycle nomogram: graphs of mathematical relationships
allow estimates of the timing of cycle events in eucaryote cells. (Eds. P ad i 11 a G. M., C a m e r o n I. L. and Z i m m e r m a n A.) In: Cell Cycle
Controls. Academic Press, New York, pp. 31-42.
K u b i t s c h e k H. E. 1970: Introduction to research with continuous cultures.
Prentice-Hall, Englewood Cliffs, New Jersey. 195 pp.
L o w y B. A. and L e i c k V. 1969: The synthesis of DNA in synchronized cultures
of Tetrahymena pyriformis GL. Expl. Cell Res., 57, 277-288.
M c D o n a 1 d B. B. 1958: Quantitative aspects of deoxyribose nucleic acid (DNA)
metabolism in an amicronucleate strain of Tetrahymena.
Biol. Bull., 114,
71-94.
M c D o n a l d B. B. 1962: Synthesis of deoxyribonucleic acid by micro- and macronuclei of Tetrahymena pyriformis. J. Cell Biol., 13, 193-203.
M i t c h i s o n J. M. 1971: The biology of the cell cycle. Cambridge University
Press, London. 313 pp.
M i t c h i s o n J. M. 1974: Sequences, pathways and timers in the cell cycle. (Eds.
P a d i l l a G. M., C a m e r o n I. L. and Z i m m e r m a n A.). In: Cell
Cycle Controls. Academic Press, New York. pp. 125-142.
M o s e r H. 1967: The mode of timing of DNA replication and of mitosis in cultured animal cells. Experientia, 23, 913-916.
N o v i c k A. and S z i l a r d L. 1950: Description of the chemostat. Science, 112,
715-716.
P e c k R. K. 1974: Morphology and morphogenesis of Pseudomicrothorax,
Glaucoma and Dexiotricha, with emphasis on the types of stomatogenesis in holotrichous ciliates. Protistologica, 10, 333-369.
P i r t S. J. 1975: Principles of microbe and cell cultivation. John Wiley and Sons,
New York. 274 pp.
P o w e l l E. O. 1956: Growth rate and generation time of bacteria, with special
reference to continuous culture. J. Gen. Microbiol., 15, 492-511.
P r e s c o t t D. M. 1957: Change in the physiological state of a cell population as
a function of culture growth and age (Tetrahymena geleii). Expl. Cell Res.,
12, 126-134.
S c h m i d t G. and T h a n n h a u s e r S. I. 1945: A method for the determination
of desoxyribonucleic acid, ribonucleic acid, and phosphoproteins in animal
tissues. J. Biol. Chem., 161, 83-89.
S m a 11 E. B. 1967: The Scuticociliatida,
a new order of the Class Ciliatea (Phylum
Protozoa, Subphylum Ciliophora). Trans. Am. Micros. Soc., 95, 739-751.
S u h r - J e s s e n P. B., S t e w a r t J. M. and R a s m u s s e n L. 1977: Timing
and regulation of nuclear and cortical events in the cell cycle of Tetrahymena pyriformis. J. Protozool., 24, 299-303.
T h o r m a r H. 1959: Delayed division in Tetrahymena
pyriformis
induced by
temperature changes. C. R. Trav. Lab. Carls., 31, 207-225.
W i 1 1 e J. J. 1972: Selective amplification of repetitive DNA in the eukaryote,
Tetrahymena.
Biochem. Biophys. Res. Comm., 46, 692-699.
W i l l i a m s N. E. 1964: Relations between temperature sensitivity and morphogenesis in Tetrahymena pyriformis GL. J. Protozool., 11, 566-572.
Z e u t h e n E. 1974: A cellular model for repetitive and free-running synchrony
in Tetrahymena
and Schizosaccharomyces.
(Eds. P a d i l l a G. M., C am e r o n I. L. and Z i m m e r m a n A.) In: Cell Cycle Controls. Academic
Press, New York. pp. 1-30.
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ACTA
Vol. 19 (No. 1)
Department
of
PROTOZOOLOGICA
WARSZAWA, 30 III 1980
Comparative Anatomy and Animal Ecology, Institute of Environmental
Biology, University of Łódź, 90-237 Łódź, Banacha 12/16, Poland
Maria
Tetratoxum
pp. 15-20
WOLSKA
unifasciculatum (Fiorent.) (Ciliata, Entodiniomorphida)
I. Somatic and Adoral Infraciliature
Received
19 April 1979
Synopsis. Silver impregnation of Tetratoxum
unifasciculatum
(Fiorent)
has revealed the occurrence of three somatic ciliary zones in this
species. The structure of the adoral zone appeared to be the same as
in Cycloposthium
Bundle and Tripalmaria
Gass. of the family Cycloposthiidae.
The genus Tetratoxum
Gass., comprising species from horse intestine, belongs to the family Ditoxidae according to S t r e l k o v (1939),
being separated from Cycloposthiidae Poche. Beside Tetratoxum the family Ditoxidae comprises the genera Ditoxum Gass., Cochliatoxum Gass.
and Triadinium Fiorent.; the last one having body shape not characteristic of the family. The base for separation of the new family by S t r e l k o v (1939) was the internal structure as well as the character of the
adoral zone, not retractable inside to the body.
Performing the systematic revision of the family Cycloposthiidae
L a t t e u r and D u f e y (1967) created the family Spirodiniidae which
embraced three subfamilies. In the subfamily Spirodininae they placed
the genus Spirodinium Fiorent (allocated by Strelkov to Ophryoscolecidae) and the genera ranged by S t r e l k o v in his family Ditoxidae, also the genus Tetratoxum Gass. These authors accepted the number and shape of ciliary zones, so called additional ones, and the presence or lack of the skeleton as the base for distinguishing of the new
family and subfamilies.
This investigation was supported by Committee of Cytobiology of the Polish
Academy of Sciences.
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16
M. W O L S K A
The above mentioned different opinions on the system of EntodiniomorphicLa from horse intestine raised up the need of further studies,
especially on the adoral ciliature of these ciliates.
Among Entodiniomorphida
from horse intestine only the infraciliature of Cycloposthiidae has been studied till now. To this family belong
two genera occurring in horse: Cycloposthium Bundle and Tripalmaria
edentatum
Gass. Detail description of infraciliature of Cycloposthium
was given by F e r n a n d e z - G a l i a n o (1959). Due to this author
it is known that the adoral zone in Cycloposthium is composed of three
clearly differring parts. The same character shows the adoral zone in
Tripalmaria dogieli examined by W o l s k a (1978). In the family Ditoxidae the species Triadinium caudatum Fiorent. has been studied by
W o l s k a (1969) but this species, being not typical of the family, is
not suitable for comparison. S e n a u d and G r a i n (1972) investigated Cochliatoxum periachtum Gass. under the electron microscope. The
protozoan was also silver impregnated but its adoral infraciliature was
not described in detail.
The aim of this work was to investigate the somatic and adoral infraciliature of the typical representative of the family
Ditoxidae,
namely Tetratoxum unijasciculatum
(Fiorent.) and to compare it with
other, already investigated representatives of Entodiniomorphida
from
horse.
The structure of T. unijasciculatum
was given by G a s s o v s k i
(1918), H s i u n g
(1930)
and
S t r e 1 k o v (1939). These authors
D.az.
Vadz
described four somatic ciliary zones in this ciliate. In the present
paper only schematic drawing of
the ciliature is given, obtained afD.adz.
Ki.c.
ter silver impregnation of the ciliates (Fig. 1).
D.p.z.
Fig. 1. Scheme of disposition of the
ciliary zones, left side view. The protozoan is represented as transparent.
Ventral side (V). Dorsal side (D). Dorsal anterior zone (D.a.z.). Dorsal posterior zone (D.p.z.). Ventral posterior
zone (V.p.z.). Ventral part of the adoral zone (V.ad.z.). Dorsal part of the
adoral zone (D.ad.z.). Kinetosomes of
free cilia (K.f.c.). Cytostome (c)
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I N F R A C I L I A T U R E OF TETRATOXVM
Material
and
UNIFASCICULATVM
17
Methods
Content of the horse colon, obtained from the sloughterhouse in Rawicz just
after killing a horse, was a source of the material. A sample of content, filtered through a gauze in order to remove larger food particles, containing protozoans was fixed in 10% formaldehyde during 24 h. After that time the protozoans
were washed in water and a drop of condensed suspension was placed on steam
heated object glass, mixed with a drop of warm gelatin (after Chatton and Lwoff)
and spread in a thin film over !the glass. Then the preparation was placed in
a moist chamber in refrigerator. Next day the preparation was submerged into
silver solution after Bielschowsky. Freshly prepared silver solution was diluted
in proportion 1 :1 with distilled water. The impregnation time was established
experimentally, usually several seconds were sufficient. After that the preparation
was quickly washed in distilled water and put into 10°/o formaldehyde till darkening. The preparation was then dehydrated and mounted in Canada balsam.
R e s u l t s
Total ciliature of Tetratoxum unifasciculatum is composed of adoral
ciliature and three somatic ciliary zones or additional zones. The somatic zones — dorsal anterior, dorsal posterior and ventral posterior
ones are in form of a ribbons. Each of these ribbons forms an arch
embracing the dorsal or the ventral margin of flattened body of the
ciliate and reaches to about the middle of the left or the right body
side (PI. I 1, Fig. 1). Each somatic zone, being rounded at both ends, is
composed of parallel rows of kinetosomes. The rows of kinetosomes are
arranged densely and evenly spaced, being diagonal to the long axis
of the zone. The kinetosomes of particular rows lay closely together, so
under a low microscope magnification they look like uniform diagonal
strips. In posterior zones almost all strips show thickenings in their
mid-length giving an appearance of a thin discontinuous band running
along the zone (PI. I 1). In the anterior (dorsal) zone two such bands
are visible (PI. 1 3).
The adoral infraciliature is differentiated into three parts. At the
ventral side there is a wide ribbon surrounding the border of an opening leading to a concavity directed toward the dorsal side and forming
the vestibulum. Narrowed endings of this ribbon, are bent inward the
concavity (Fig. 1). This ventral part of the adoral infraciliature is composed of dense, parallel and evenly arranged diagonal rows of kinetosomes, being similar to the somatic zones. Along this ribbon, as in the
dorsal somatic zone, two dark bands are visible (PI. I 3). The second part
of the adoral infraciliature is in form of a narrow ribbon forming a loop
2 — Acta Protozool. 1/80
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18
M. W O L S K A
directed postero-dorsally. The endings of the dorsal ribbon butt against
the endings of the ventral one. A narrow ribbon, forming the dorsal part
of the adoral infraciliature is composed of short rows of kinetosomes.
The kinetosomal rows are more loosely arranged than in the ventral
part and particular kinetosomes are more intensely blackned (PI. II 4).
The dorsal ribbon of short rows runs in waves on the bottom of vestibulum
surrounding a slit-like cytostome. The third part of the adoral ciliature constitutes a group of kinetosomes lying on the ventral side of
the ciliate body and being slightly shifted to the right, near the posterior margin of the ventral part of adoral ciliature (Fig. 1, PI. I 2,
II 5).
Discussion
The ciliary zones in T. unifasciculatum,
recognized after silver impregnation, show the same character as the ciliary zones of up to
now examined Entodiniomorphida
from horse intestine and cattle rumen. All zones are composed of diagonal rows of kinetosomes, parallel
and situated in the same distances. Such arrangement of kinetosomal
rows in ciliary zones was recognized by N o i r o t - T i m o t h e e (1960)
in Ophryoscolecidae from cattle rumen and, somewhat earlier, b y F e r n a n d e z - G a l i a n o (1959) in Cycloposthium edentatum (fam. Cycloposthiidae) from horse. Recently, the same pattern of ciliary rows has
been found in another representative of Cycloposthiidae, namely in
Tripalmaria dogieli by W o 1 s k a (1978).
In T. unifasciculatum
the membranellae are lacking in somatic zones as well as in the adoral one although the cilia in these zones are
grouped into larger units, the syncilia, similarly as in other Entodiniomorphida. The adoral zone of T. unifasciculatum
shows no particularities; its structure fits well the structure of the adoral zone characteristic of Cycloposthiidae. It may be characterized by different arrangement of kinetosomal rows and more intense impregnation of kinetosomes in the dorsal than in the ventral part. Such difference in structure between the ventral and the dorsal parts does not exist in Ophryoscolecidae. At the posterior border of the ventral part of adoral zone
in T. unifasciculatum occurs a group of kinetosomes pertaining to a prominent cytoplasmic lip surrounding the adoral zone. Presence of this
group of kinetosomes (free cilia kinetosomes) is a common character
of Cycloposthiidae and Ophryoscolecidae
(Fernandez-Galiano
1959, W o l s k a 1965, 1978). In comparison with Cycloposthiidae the
dorsal part of the adoral zone in T. unifasciculatum is more strongly
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I N F R A C I L I A T U R E OF TETRATOXUM
UNIFASCICULATVM
19
developed and, what is more, the disjunction between the dorsal and
the ventral parts, well visible in Cycloposthiidae, is not visible in
this species. The dorsal and the ventral parts of the adoral zone develop from two separate primordia in Cycloposthiidae and Ophryscolecidae ( N o i r o t - T i m o t h e e 1960, W o l s k a 1965). As T. unifasciculatum is concerned, the protozoans in course of division have not
been found in the examined material, so the observations on the number of adoral zone primordia could not be made. In order to elucidate
this problem further investigations are necessary as well as more comprehensive examination of this species under discussion with the aid
of electron microscope.
RÉSUMÉ
L'imprégnation de Tetratoxum
unifasciculatum
(Fiorent.) à l'argent démontre
la présence de trois zones ciliaires somatiques. La zone adorale se caractérise par
la même composition que chez Cycloposthium
Bundle et Tripalmaria
Gass. de
la famille des
Cycloposthiidae.
REFERENCES
F e r n a n d e z - G a l i a n o D i m a s 1959: La infraciliation en Cycloposthium
edentatum Strelkov. Boln. R. Soc. Esp. Hist. Nat., 57, 139-150.
G a s s o v s k y G. 1918: K mikrofaunie kiSeCnika loSadi. Trudy petrogr. Ob§£.
Estest., 49, 30-36.
H s i u n g T. S. 1930: A monograph on the Protozoa of the large intestine of the
horse. Iowa St. Coll. J. Sci., 4, 359-423.
L a t t e u r B . et D u f e y M . M. 1967: Reforme systématique de la famille des
Cycloposthiidae
Poche, 1913. Acta Zool. Pathol. Antverpiensia, 44, 125-139.
N o i r o t - T i m o t h é e C. 1960: Etude d'une famille de ciliés: Les
"Ophryoscolecidae". Structure et ultrastructure. Annls. Sci. Nat. Zool., 12 serie, 527-718.
S e n a u d J . et G r a i n J. 1972: Etude ultrastructurale préliminaire de Cochliatoxum periachtum
Gassovsky, 1919, cilié Entodiniomorphe endocommensal
du cheval. Protistologica, 8, 65-82.
S t r e l k o v A. 1939: Paraziticeskie infuzorii iz kiâeënika neparnokopytnych sem.
Equidae. Uc. zap. Leningr. Gos. Pedagog. Inst. Im. A. I. Gercena, 17, 1-262.
W o l s k a M. 1965: Remarks on the adoral ciliature in the order
Entodiniomorpha.
Acta Protozool. 3, 321-325.
W o l s k a M. 1969: Triadinium minimum Gassovsky — its phylogenetic significance.
Rev. Soc. Mex. Hist. Nat. 30, 65-78.
W o l s k a M. 1978: Tripalmaria
dogieli Gass., 1918 (Ciliata,
Entodiniomorphida).
Structure and ultrastructure. Part I. Light-microscope investigations. Acta
Protozool., 17, 13-20.
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EXPLANATION OF PLATES I-II
Tetratoxum unifasciculatum,
photographs of silver impregnated specimens
1: A general view. Dorsal anterior zone (D.a.z.). Dorsal posterior zone
Ventral posterior zone (V.p.z.). Adorai zone (Ad.z). X 700
2: Ventral part of adorai zone (V.ad.z.). Kinetosomes of free cilia (arrow.)
3: Dorsal anterior zone (D.a.z.). Ventral part of the adorai zone (V.ad.z.).
part of the adorai zone (D. ad.z.). X 1700
4: Dorsal anterior zone (D.a.z.). Dorsal part of the adorai zone (D.ad.z.).
5: Dorsal anterior zone (D.a.z.). Ventral part of the adorai zone (V.ad.z.).
part of the adorai zone (D.ad.z.). Kinetosomes of free cilia (arrow). X 1700
http://rcin.org.pl
(D.p.z.).
X 1700
Dorsal
X 1700
Dorsal
ACTA
PROTOZOOL.
VOL.
19, N o .
1
PLATE
D.O.Z.
* > ' Vad.z.
V p . z . 7
D p . z .
i
M. Wolska
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I
A C T A P R O T O Z O O L . V O L . 19, N o . 1PLATEVII
D.a.z.
V.ad.z.
\
D.acf.z
M. Wolska
auctor phot.
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ACTA
Vol. 19 (No. 1)
Department
of
PROTOZOOLOGICA
WARSZAWA, 30 III 1980
pp. 21-28
Comparative Anatomy, and Animal Ecology, Institute of Environmental
Biology, University of Łódź, 90-237 Łódź, Banacha 12/16, Poland
Maria W O L S K A
Tetratoxum
unijasciculatum (Fiorent.) (Ciliata,
Entodiniomorphida)
II. Electron Microscope Investigations
Received
19 April 1979
Synopsis. The study has been performed on ultrastructure of the cortex
and of the buccal apparatus of Tetratoxum unijasciculatum.
It has been
found that under cell membrane and epiplasm of bare parts of the body
there is a layer of longitudinal microtubules arranged in bundles under
which longitudinal bars of dense substance are situated. Short kinetosomes are dispersed in microfibrillar layer occurring under these bars.
The kinetosomes of the ciliary zones are not grouped although the cilia
join together and form syncilia. Under kinetosomes of ciliary zones
spreads a net of dense substance in which strands parallel to kinetosomal rows are dominant. The structure of T. unifasciculatum
cortex is
similar to that of Triadinium caudatum and Cochliatoxum
periachtum
of the same family. Also the complex cytostome-cytopharynx shows the
same character as in both mentioned species. A group of cilia called
"free cilia" occurs also in T. unifasciculatum;
their ultrastructure being
the same as in other examined representatives of
Entodiniomorphida
and
Blepharocorythina.
New concepts of the system of Ciliata have changed the position
of Entodiniomorphida
( d e P u y t o r a c e t a l . 1974, C o r l i s s 1975,
1977, J a n k o w s k i 1973, S e r a v i n and G e r a s s i m o v a 1978).
The opinions are concordant that Entodiniomorphida
ought to be removed from former class Spirotricha and placed in more primitive one,
but the rank of this group is still discutable as well as its supposed
relations with Blepharocorythina.
Also the system within Entodiniomorphida from horse intestine is differently considered by various authors ( W o l s k a 1971). Thus, the need arises of more extensive studies
on these ciliates. Recently, Tetratoxum unijasciculatum,
the representative of the family Ditoxidae created by S t r e l k o v (1939), has been
examined using silver impregnation method (W o 1 s k a 1980). The aim
of the present paper is to recognize the ultrastructure of the cortex
and of the buccal apparatus of this species.
This investigation was supported by Committee of Cytobiology of the Polish
Academy of Sciences.
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22
Material
and
Methods
The material was taken from the content of horse colon in Rawicz sloughterhouse. Samples of content were taken just after killing the horse and put in
vacuum flasks. Just after bringing to the laboratory the protozoans from intestinal content were prepared for embedding in Epon 812 according to the method of
G r a i n (1966). Sections cut on ultramicrotome III LKB were placed on Formvarcoated grids and contrasted with uranyl acetate and lead citrate. Observations
were made in Tesla BS 500 electron microscope. Semi-thin sections were made
according to the procedure described earlier (W o 1 s k a 1978 a).
R e s u l t s
The body of Tetratoxum unifasciculatum
is covered by single cell
membrane spreading also over the cilia. Under this membrane in nonciliated regions there is a thin layer of structureless substance of medium electron density — the epiplasma (PI. I 1, II 5). Somewhat deeper
longitudinal bundles of microtubules are situated. In most cases the
bundle comprises six microtubules lying at two levels by three (PI. I 1,
II 5). Among bundles of microtubules there are vesicles comprising granular content or empty ones (PI. I 3, II 4), some of them being opened
outside (PI. I 1). Under the bundles of microtubules there are longitudinal bars of dense substance taking exactly the same course as the
microtubules (PI. I 1, 3, II 4, 6). The longitudinal bars are connected
by thin transverse strands of dense substance (PI. I 1, 3, III 7). The longitudinal bars are underlayered by circular microfibrils (PI. I 1, 2, II 4)
with short nonciliated kinetosomes being dispersed among them (PI. I
2, 3, II 4, 6). Moreover, the microfibrils proliferate the whole thickness
of the ectoplast which, what is known in Ditoxidae ( S t r e l k o v 1939),
does not surround the endoplasmic sac from all sides being limited
to the anterior part of the body and partly to the right side. A compact
strand of microfibrils occurs in the region of surface ribs on the ventral and on the dorsal body sides. This strand runs circularly at the
base of ribs (PL III 8). The microfibrils delimit also the ectoplast from
endoplast (endoplasmic sac) forming an inconspicuous layer here and
there (PI. Ill 7, 8), or well visible double layer (PI. Ill 9). Bars of dense
substance do not occur in cytoplasmic lips surrounding the zones of
cilia and in the vault of vestibulum (PI. Ill 10).
The ciliary zones, in form of ribbons, are composed of parallel evenly spaced rows of cilia (PI. IV 12, 14). It has been already shown on
silver impregnated total preparations (W o 1 s k a 1980). The kinetosomes
have opened bases (PI. IV 13). Structureless dense substance at the
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U L T R A S T R U C T U R E OF TETRATOXVM
UNIFASCICULATUM
23
base of kinetosomes slightly narrows lumen of the kinetosome cylinder
and gives branchings to inside of the cell and to neighbouring kinetosomes. As a result, a complicated spatial net is formed spreading beneath
the ciliary zones. In this intricate dense net the most pronounced are the
strands running deeply along the rows of kinetosomes and being connected by transverse commissures (PI. IV 14, VI 17 a). Derivative fibres
get off from kinetosomes. Some of them are microtubular transverse
fibres, the other ones are supposed to be feebly developed kinetodesmal fibrils (PI. IV 14). The nematodesms, abundant in T. unijasciculatum, arise from the dense net. Some nematodesm are large and spread
far behind the ciliary zones (PI. IV 11, V 15). In places of large nematodesms origin dense substance of the net forms greater aggregations
(PI. IV 17) corresponding to the thickening of kinetosomal rows revealed in silver impregnated preparations ( W o l s k a 1980). The nematodesm are especially numerous in the adoral zone (PI. V 16). They form
a timbering of cytopharynx in their further course. The cytoplasmic
lips surrounding the ciliary zones are bent in their distal parts and
form a characteristic pattern in sections (PL IV 12, X 28).
The cilia in T. unijasciculatum are joined into groups, similarly as
in other Entodiniomorphida.
For these groups the name syncilia has
been admitted. S e n a u d and G r a i n (1972) have observed that in
Cochliatoxum periachtum the axis of syncilium does not coincide with
antero-posterior axes of kineties. The same phenomenon may be observed in T. unijasciculatum (PL IV 12).
At the ventral margin of the protozoan body, within the area of
adoral zone lip, the cytoplasma is deeply ploughed and short deformed cilia are embedded in furrows (Pl. VI 18, VII 19, IX 25). In folds
between furrows a layer of microtubular fibres occurs under cell membrane (Pl. VII 20). The kinetosomes of mentioned cilia (free cilia) have
been already revealed in T. unijasciculatum
by silver impregnation
method ( W o l s k a 1980). These cilia and folds occupy a small area in
shape of a belt beginning at the right side of the body, overpassing
ventral margin and spreading over the left side (cf. W o l s k a 1980).
Left extremity of this structure is slightly bent backward and enters
deeply the base of the adoral zone lip. So, in some sections this structure may be cut twice (Pl. VI 18, IX 25). The cilia are swollen or flattened and so short that they do not reach even the half height of the
folds (Pl. VII 21). The axosomes are lacking in kinetosomes of this
group of cilia but septum and axial granules are present (Pl. VII 19).
The pattern of microtubular fibres is disturbed in most cilia. Most frequently the axial fibres can not be distinguished and the number of
pairs of peripheric fibres differs from nine (Pl. VII 22 a). In some cilia
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24
M. W O L S K A
fibres are crowded disorderly (PI. VII 22). Some cilia of this group,
situated peripherically, retain normal pattern of fibres.
The cytostome is situated in the wall of a concavity (vestibulum) the
inlet of which is surrounded by the ventral part of the adoral zone
( W o l s k a 1980). In general the vestibulum is directed slightly to the
back and to the dorsal right side of the body, but its particular segments deviate from this direction. The cytoplasm around the cytostome
is slightly raised forming a rampart covered by short rows of cilia of
the dorsal part of the adoral zone (PI. VIII 23). Wide cytopharynx, folded in transverse and longitudinal sections, is directed to the dorsal back
and to the right side (PI. VIII 24, IX 26, 27). Along the vault of the
vestibulum run the folds similar to cytoplasmic lips surrounding somatic
ciliary zones (PI. X 28, 30). The cytostome is provided with microtubules
(rideaux de tubules) derived from the cilia neighbouring the cytostome
(PI. X 29). The wall of cytopharynx is composed of the layer of nematodesms, small groups of microtubules and a compact fibro-granular layer
(PI. XI 31, 32, XII 33). Outside to it there are numerous large nematodesms (PI. XI 31, XII 33) originating from the dense net underlying
the adoral ciliary zone. The microfibrils are interlaced among these
nematodesms (PI. XI 31, XII 33).
Discussion
Comparison of structure of Cycloposthiidae sensu lato examined under the electron microscope has shown that the cortex of T. unifasciculatum is the most similar to that of Triadinium caudatum ( W o l s k a
1978 b). In nonciliated parts of the body under cell membrane in both
species there is a thin layer of epiplasm, then the layer of microtubules
arranged in bundles. Under them there are the bars od dense substance
and finally the microfibrillar layer with short nonciliated kinetosomes
dispersed without any order. Moreover, at the level of microtubules and
partly at the level of bars, numerous vesicles are present in both species. Probably they secrete mucus visible sometimes on the cell membrane as a fuzz. The cortex of T. unifasciculatum differs only slightly
from Cochliatoxum periachtum (S e n a u d and G r a i n 1972) by the
presence of epiplasma layer and another shape of elements of dense
substance situated between microtubular and microfibrillar layers. In
C. periachtum they are in form of strongly developed plates instead of
bars.
In Tripalmaria dogieli ( W o l s k a 1978 a) all mentioned elements of
the cortex are arranged in the same order but in addition to that there
is a polysaccharide skeleton embraced by microfibrillar layer at the
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U L T R A S T R U C T U R E OF TETRATOXVM
UNIFASCICULATUM
25
right side of the body. Such skeleton is lacking in T. unifasciculatum as
well as in T. caudatum and C. periachtum. T. unifasciculatum is the
most different from Cycloposthium bipalmatum ( G r a i n 1966). The latter does not show distinct splitting of microtubular layer in bundles
or the dense substance in form of bars or plates. In C. bipalmatum
the mucus vesicles do not occur but there is a polysaccharide skeleton
in almost the whole body between the microtubular and microfibrillar
layers. The kinetosomes in nonciliated body parts of C. bipalmatum
have not been recognized.
Organization of the ciliary zones in T. unijasciculatum is the same
as in other Entodiniomorphida.
The cilia are grouped in syncilia, while
in the level of kinetosomes grouping are lacking ( N o i r o t - T i m o t h e e 1960, G r a i n 1966, S e n a u d and G r a i n 1972). The kinetosome cylinder in T. unijasciculatum is open similarly as in T. caudatum
and C. periachtum, being thus different from T. dogieli and C. bipalmatum. It seems that the kinetosomes of T. unifasciculatum have the same
associated fibres as C. periachtum and T. caudatum, namely transverse
fibres and kinetodesmal ones. All kinetosomes of ciliary zones in T.
unifasciculatum are included into a common net of dense substance formed by ramification of "manchets" surrounding the bases of kinetosomes.
It is very similar to the arrangement of dense substance joining the
kinetosomes in T. caudatum and C. periachtum. The connections between
kinetosomes exist also in other Entodiniomorphida but dense substance
has another form. The cytostome-cytopharynx complex in T. unifasciculatum has the same character as in C. periachtum and T. caudatum.
The cytopharyngeal wall is composed of the layer of microtubules arranged in small groups and of the outer compact granular layer. This
wall is folded and surrounded by nematodesms. Another structure has
the wall of cytopharynx in C. tripalmaria.
In sum, the ultrastructure of the cortex and of the buccal apparatus
in T. unifasciculatum,
C. periachtum and T. caudatum is very similar.
The genera Tetratoxum, Cochliatoxum and Triadinium, as well as Ditoxum, characterized by lack of skeleton, non retractible adoral zone
and reduced ectoplast, have been taken out by S t r e l k o v (1939) from
the family Cycloposthiidae Poche and placed in a new family. The results of electron microscope investigation of the representatives of three
of these genera uphold Strelkov's opinion. Simultaneously S t r e l k o v
(1939) transferred the genus Spirodinium Fiorent from the old family
Cycloposthiidae to Ophryoscolecidae forming a new subfamily Spirodiniinae for it. S t r e l k o v quotes some morphological characters of
this genus approaching it to Ophryoscolecidae. Another opinion has been
pronounced by L a t t e u r and D u f e y (1967). These authors relate
Spirodinium
to Tetratoxum,
Cochliatoxum,
Triadinium and Ditoxum
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26
M. W O L S K A
comprising all these genera in the subfamily Spirodiniinae within the
new family Spirodiniidae. Further investigations on Spirodinium,
in
light as well as in electrons microscope, should give answer whether
this genus ought to be placed within the family Ophryoscolecidae or
within the distinct family Spirodiniidae (subfamily Spirodiniinae) together with Tetratoxum, Cochliatoxum, Triadinium and Ditoxum.
The "free cilia" in T. unifasciculatum have the same ultrastructure
as in Triadinium caudatum (W o l s k a 1978 b) and Tripalmaria dogieli
(W o 1 s k a 1978 a), as well as in the representatives of Blepharocorythina — Ochoterenaia appendiculata (W o l s k a 1978 c) and Circodinium
minimum (W o 1 s k a 1979). Thus, "free cilia" are common structures of
Entodiniomorphida
and Blepharocorythina.
Such generalization is justified because the kinetosomes of "free cilia" have been recognized in
light microscope also in other representatives of
Entodiniomorphida
( F e r n a n d e z - G a l i a n o 1959, W o l s k a 1965) and
Blepharocorythina from horse intestine (W o l s k a 1971) beside the species mentioned in the present paper.
REFERENCES
C o r l i s s J. O. 1975: Taxonomic characterization of (the suprafamilial groups in
a revision of recently proposed schemes of classification for the phylum
Ciliophora. Trans. Am. Microsc. Soc., 94, 224-267.
C o r l i s s J. O. 1977: Annotated assignment of families and genera to the orders
and classes currently comprising the corlissian scheme of higher classification for ithe phylum Ciliophora. Trans. Am. Microsc. Soc., 96, 104-140.
F e r n a n d e z - G a l i a n o D i m a s 1959: La infraciliation en Cycloposthium
edentatum Strelkov. Bol. R. Soc. Esp. Hist. Nat. Secc. Biol., 57, 139-150.
G r a i n J. 1966: Etude cytologique de quelques Ciliés Holotriches endocommensaux des Ruminants et des Equides. Thèse, Clermont-Ferrand éditions du
C.N.R.S., 1-141.
J a n k o w s k i A. W. 1973: Taxonomic revision of subphylum Ciliophora Doflein,
1901. Zool. Zh. 52-175.
L a t t e u r B. et D u f e y M. M. 1967: Réforme systématique de la famille des
Cycloposthiidae
Poche, 1913. Acta Zool. Pathol. Antverpiensia, 44, 125-139.
OphryoscoleN o i r o t - T i m o t h é e C. 1960: Etude d'une famille de Ciliés: les
cidae. Structures et ultrastructures. Annls. Sci. Nat. Zool., 2, 527-718.
De
P u y t o r a c P., B â t i s s e A., B o h a t i e r J., C o r l i s s J. O.,
Der o u x G., D i d i e r P., D r a g e s c o J . ,
Fryd-VersavelG.,
Grain
J., G r o l i e r e C. A., H o v a s s e R., I f t o d e F., L a v a l M., R o q u e
M., S a v o i e A. et T u f f r a u M. 1974: Proposition d'une classification
du phylum Ciliophora Doflein, 1901 (réunion de systématique, ClermontFerrand). C. R. Acad. Sci., Paris, 278, 2799-2802.
S e n a u d J . et G r a i n J. 1972: Etude ultrastructurale préliminaire de Cochliatoxum periachtum Gassovsky, 1919, cilié endocommensal du cheval. Protistol o g y , 8, 65-82.
S e r a v i n L. N. and G a r a s s i m o v a Z. P. 1978: A new macrosystem of Ciliates. Acta Protozool., 17, 399-418.
S t r e l k o v A. 1939: Parasiticeskie infuzorii iz kiëefnika neparnokopytnych semejstwa Equidae. Uë. Zap. Leningr. Gos. Pedagog. Inst. Im. A. I. Gercena,
17, 1-262.
W o l s k a M. 1965: Remarks on the adoral ciliature in the order
Entodinimorpha.
Aota Protozool., 3, 321-325.
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U L T R A S T R U C T U R E O F TETRATOXVM
UNIFASCICULATUM
27
W o l s k a M. 1971: Studies on the family Blepharocorythidae
Hsiung. V. A review
of genera and species. Acta Protozool. 9, 23-40.
W o l s k a M. 1978a: Tripalmaria
dogieli Gass, 1918 (Ciliata,
Entodiniomorphida).
Structure and ultrastructure. Part II. Electron-microscope investigations.
Acta Protozool. 17, 21-30.
W o l s k a M. 1978 b: Triadinium caudatum Fiorent. Electron Microscope Examinations. Acta Protozool. 17, 445-454.
W o l s k a M. 1978 c: Light and electron microscope studies on Ochoterenaia
appendiculata
Chavarria (Ciliata, Blepharocorythina).
Acta Protozool. 17, 483492.
W o l s k a M. 1979: Circodinium minimum (Gassovsky, 1918), (Ciliata, Blepharocorythnina). Electron microscope investigation. Acta Protozool. 18, 223-229.
W o l s k a M. 1980: Tetratoxum
unifasciculatum
(Fiorent), (Ciliata,
Entodiniomorphida). I. Somatic and adorai infraciliature. Acta Protozool. (in press).
RÉSUMÉ
L'ultrastrueture du cortex et de l'appareil oral a été étudiée chez le Tetratoxum unifasciculatum
(Fiorent). Dans les parties du corps dépourvues de la ciliature on trouve, sous la membrane cellulaire et l'épiplasme, une couche des
microtubules longitudinales disposées par faisceaux et, plus profondement, des baguettes longitudinales d'une substance dense. Au dessous de ce système caractéristique des baquettes les courts cinétosomes sont dispersés dans une couche microfibrillaire. Dans les zones ciliés les cinétosomes ne sont pas groupés en dépit du
fait que les cils fusent en formant des syncils. Sous les cinétosomes des zones
ciliés s'étend un réseau composé d'une substance dense avec la prédominance des
bandes dirigées en parallèle par rapport aux rangs des cinétosomes. La structura
du cortex chez T. unifasciculatum
est pareille à celle du Triadinium caudatum et
du Cochliatoxum
periachtum,
appartenants à la même famille. Le complexe cytostome-cytopharynx porte également le même caractère chez ces trois espèces. La
groupe des cils appelles "free cilia" est présent chez T. unifasciculatum.
Us ont
la même ultrastructure que chez les autres Entodiniomorpha
et
Blepharocorythina
étudiés jusqu'à présent.
EXPLANATION OF PLATES I—XII
Tetratoxum
unifasciculatum
(Fiorent.), l-22a and 28-33 — electronograms, 23-27 —
microphotographs of semi-thin sections.
1: Transverse section perpendicular to body surface. Cell membrane (M), epiplasm (Ep), microtubules (Tu), vesicles (V), longitudinal bars (Lb), transverse
strands (Ts), microfibrils (Mf). X 35 000
2: Section oblique to body surface. Microfibrils Mf), kinetosome (K). X 36 400
3: Section tangent to body surface, slightly oblique. Vesicles (V) with granular
content or empty ones seen among microtubules, kinetosomes (K) dispersed in
microfibrillar layer; transverse strand (Ts) of dense substance. X 23 000
4. Oblique section. Vesicles (V) among longitudinal microtubules; microfibrillar
layer (Mf), kinetome (K). X 32 000
5: Transverse section. Microtubules (Tu), epiplasm (Ep), X 39 000
6: Section tangential to body surface. Vesicles (V), longitudinal bars (Lb), microfibrils (Mf), kinetosome (K). X 18 400
7: Transverse section. Transverse strands of dense substance (Ts); delimination
of ectoplast from endoplasmic sac is seen. X26 000
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8: Transverse section through the region of ribs in the anterior body part. Strand
of microfibrils (Mf) at the base of ribs. X 23 000
9: Microfibrils at the boundary of ectoplast and endoplasmic sac. Nematodesms
in ectoplast (Nd). X 41 400
10: Section through the vault of vestibulum. X 39 000
11: Section through ectoplast. Numerous nematodesms. X 13 800
12: Section through somatic ciliary zone. Groups of cilia and direction of kineties
is seen. X 6700
13: Longitudinal section through kinetosomes. Open bases of kinetosomes are seen.
Strand of dense substance (arrow). X 19 200
14: Transverse section, slightly oblique, through kinetosomes of ciliary zone. Kinetodesmal fibres (Kd), transverse fibres (T). Double strands of dense substancerunning under kineties and transverse strands are visible. X 32 000
15: Large nematodesma originating from dense substance net. X 19 200
16: Section through adoral zone. Abundance of nematosms X 19 200
17: Mass of dense substance in the place of origin of nematodesma (arrow).
X33 600
17 a: Longitudinal section through kinetosomes. Mass of dense substance going
to inside and to neighbouring kinetosomes (arrow). X 32 200
18: Section through "free cilia" region. Twice cut folds and cross-sections of some
cilia are seen. X 19 200
19: Section through folds and "free cilia" near adoral zone. Adoral zone cilia
(Ad.c.), kinetosomes of "free cilia" (K.f.c.) cytoplasmic lip of the adoral zone
(C.I.). X 26 000
20: Section through folds separating "free cilia". Microtubules (arrow). X 33 600
21: Section through folds and deformed "free cilia". A very short cilium (arrow).
X 24 000
22: Section through deformed cilia. Atypical pattern of fibres in cilia. X 49 000
22 a: The same. Cross-section through a cilium with 6 pairs of tubular fibres
(arrow). X 49 000
23: Section in plain of cilia surrounding cytostome. Ventral side (V), dorsal side
(D), X 1700
24: Section similar to the preceding nearer right side of the body. Ventral side
(V), dorsal side (D), section of cytopharynx is visible. X 1700
25: Approximately sagittal section near left body side. "Free cilia" (arrow), ventral side (V), X 1700
26: Longitudinal oblique section near right body side. Cytopharynx directed postero-dorsally. Macronucleus is seen. Dorso-right side (D.R.). X 1700
27: Section similar to above one. Cilia of the anterior dorsal somatic zone (arrow).
X 1700
28: Section through cytoplasmic lip of the somatic ciliary zone. X 19 200
29: Section through kinetosomes and originating from them microtubular fibres
forming an equipment of cytostome. X 20 000
30: Section through vestibulum and cytopharynx. X 15 000
31: Section through vestibulum and cytopharynx. Outer nematodesms (arrow),
microfibrill (Mf). X 10 000
32: Section through cytopharynx. Groups of microtubules (Tu), fibro-granular
layer (arrow). X 19 200
33: Section through vestibulum and cytopharynx. Nematodesms and small groups
of microtubules (arrow), outer nematodesms (Nd), microfibrils (Mf), "Rideaux de
tubules" (R). X 28 000
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ACTA PROTOZOOL. VOL. 33, N o . 1
PLATE I
f t
M. Wolska
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I ' l l
auctor phot,
A C T A P R O T O Z O O L . VOL. 19, N o . 1
M. Wolska
P L A T E VII
auctor phot.
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ACTA P R O T O Z O O L . VOL. 19, N o . 1
P L A T E III
>
1
0
M. Wolska
auctor phot,
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A C T A P R O T O Z O O L . V O L . 19, N o . 1
P L A T E VII
auctor phot.
M. Wolska
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P L A T E VII
A C T A P R O T O Z O O L . V O L . 19, N o . 1
auctor phot.
M. Wolska
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A C T A P R O T O Z O O L . VOL. 19, N o . 1
M. Wolska
P L A T E VI
auctor phot.
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P L A T E VII
A C T A PROTOZOOL. V O L . 19, N o . 1
auctor phot.
M. Wolska
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A C T A P R O T O Z O O L . V O L . 19, N o . 1
M. Wolska
P L A T E VIII
auctor phot.
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P L A T E VII
ACTA P R O T O Z O O L . VOL. 19, N o . 1
auctor phot.
M. Wolska
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A C T A P R O T O Z O O L . VOL. 19, N o . 1
M. Wolska
P L A T E VIII
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ACTA
PROTOZOOL.
VOL.
19,
Ńo.
1
M. Wolska
PLATE
xl
auctor phot.
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M. Wolska
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ACTA
Vol 19 (No. 1)
Zooogisches
Institut
PROTOZOOLOGICA
WARSZAWA, 30 III 1980
der Universität
Salzburg,
Wilhelm
Akademiestr,
pp. 29-50
26 A-5020 Salzburg,
Austria
FOISSNER
Taxonomische Studien über die Ciliaten des Großglocknergebietes
(Hohe Tauern, Österreich). VI. Familien Woodruffiidae,
Colpodidae und Marynidae
Received
12 July 1979
Synopsis. Es wird die Morphologie, die Infraciliatur und das Silberliniensystem einiger neuer oder wenig bekannter Colpodida aus den Kleingewässern des Großglocknergebietes (Hohe Tauern, Österreich) beschrieben. Folgende neue Taxa werden errichtet: Platyophrya
citrina nov.
spec., Platyophrya
dubia nov. spec., Platyophrya
hyalina nov. spec.,
Platyophrya
procera nov. spec., Colpoda ovinucleata
nov. spec., Colpoda rotunda nov. spec. und Colpoda variabilis nov. spec. Die Infraciliatur und das Silberliniensystem von Maryna ovata weisen darauf
hin, daß die Marynidae mit den Colpodidae eng verwandt sind.
In einer früheren Studie (F o i s s n e r 1978) habe ich mich eingehend
mit der Systematik der Colpodida auseinandergesetzt und die Ordnung
in zwei Unterordnungen aufgeteilt. Während der Untersuchungen über
die Ciliatenfauna der Kleingewässer des Großglocknergebietes fand ich
mehrere neue Colpodida, deren Morphologie und Infraciliatur Gegenstand dieser Arbeit ist. Für die schon damals hervorgehobene enge
Verwandtschaft der Gattungen Platyophrya und Cyrtolophosis können
nun weitere Belege beigebracht werden. Bei Platyophrya hyalina ist
der Mund so wie bei Cyrtolophosis mucicola deutlich auf die Ventralseite verlagert und etwas eingesenkt. Vor kurzem fand ich in alpinen
Böder. außerdem eine Cyrtolophosis verwandte Art, bei der die linke
Körperseite fast ganz unbewimpert ist, ähnlich wie bei der Gattung
Platyophrya (s. F o i s s n e r 1978). Im gleichen Biotop entdeckte ich
weitere neue Species der Genera Platyophrya und Cyrtolophosis. Ihre
Beschreibung wird an anderer Stelle veröffentlicht. Diese Beobachtunge.i weisen darauf hin, daß ein Großteil der Arten der Familien
Wood'uffiidae und Cyrtolophosididae noch unbekannt ist.
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30
W. F O I S S N E R
Material,
Methoden,
Abkürzungen
Das Untersuchungsmaterial wurde in den Kleingewässern (Weidetümpeln, ausgedrückten Moosen von kleinen Bächen etc.) entlang der Großglockner-Hochalpenstraße gesammelt. Genauere Fundortangaben und ökologische Daten finden sich
bei F o i s s n e r (1980). Darauf bezieht sich auch die Numerierung des Kleingewässers beim Locus typicus der neuen Arten.
Zur Darstellung der Infraciliatur und des Silberliniensystems verwendete ich
die nasse Silberimprägnationsmethode von C o r I i s s (1953) und das trockene
Silberimprägnationsverfahren von F o i s s n e r (1976). Der Kernapparat wurde
mit Orcein-Essigsäure, die Protrichocysten wurden mit Methylgrün-Pyronin (MP)
angefärbt ( F o i s s n e r 1979). Alle Arten wurden einer sorgfältigen Lebendbeobachtung unterzogen.
A
aM
Ba
CV
D
d
Dr
DV
E
K
G
Gefressene Grünalgen
Adorale Membranellen
Bakterien (?)
Kontraktile Vakuole
Großer Dorsalsack
Kleiner Dorsalsack
Diagonalrinne
Defäkationsvakuole
Kompakte Entoplasmaeinschlüsse
— Gallertgehäuse
— Kristalle
—
—
—
—
—
—
—
—
—
1P
mK
NV
P
pM
Pt
rP
T
V
Vk
Vn
z
—
—
—
—
—
—
—
—
—
—
—
—
Linke Polykinete
Mycterale Kineten
Nahrungsvakuole
Exkretionsporus der CV
Parorale Membran
Protrichocysten
Rechte Polykinete
Trichiten
Vestibulum
Vestibularkineten
Ventralnaht am Kiel
Zoochlorellen
Beschreibung der Arten
Außer den unten beschriebenen Arten fand ich noch Cyrtolophosis
mucicola Stokes, Colpoda cucullus Müller, Colpoda inflata (Stokes) und
Colpoda steinii Maupas. Den Beschreibungen dieser Species durch K a h l
(1930-35), F o i s s n e r (1978) u.a. habe ich nichts hinzuzufügen.
1. Familie Woodrujfiidae
Platyophrya
Gelei
citrina nov. spec. (Abb. 1 a-d)
D i a g n o s e : 40-60 |im große, etwa 2 : 1 abgeflachte, stark metabolische Platyophrya, deren Entoplasma leicht gelborange gefärbt ist.
Ventrallinie sigmoid, Dorsallinie im proximalen Abschnitt meist stark
gewölbt, so daß der Mund weit nach links verschoben wird. Rechts
7-9 Kineten mit je etwa 30 Basalkörperpaaren. Links 6-7 Kineten mit
je ca. 12 Basalkörperpaaren, einschließlich der dicht unterhalb der
4-5 kleinen adoralen Membranellen liegenden Dikineten.
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CILIATEN DES
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31
L o c u s t y p i c u s : Vereinzelt in einer tümpelartigen Verbreiterung eines reinen Bächleins (Tümpel 27) südlich des Fuschertörls (Großglockner-Hochalpenstraße, etwa 2300 m ü.d.M.).
M o r p h o l o g i e : Körperform variabel, aber sehr charakteristisch
(Abb. 1 a, d). Unterschiedlich ist vor allem die Stärke der Wölbung der
Dorsallinie. Körperquerschnitt proximal lang oval, distal kurz oval
(Abb. 1 b, c). Mund von sehr zarten Trichiten umgeben. Makronucleus
kugelförmig, mit länglichen Nucleolen und dicht anliegendem Mikronucleus. Dicht unter der Pellicula viele leicht zylindroide Protrichocysten (Abb. 1 a). Sie werden in Form von kleinen Tröpfchen abgesondert, die rasch zu einem umfangreichen, feinmaschigen Netzwerk verquellen ( F o i s s n e r 1979). Kontraktile Vakuole subterminal, mit kurzem, breitem Röhrenporus. An der Stelle seiner Ausmündung ist das
Tier stets leicht eingedellt. Entoplasma mit vielen, oft sehr großen, orangegrünen Einschlüssen (Abb. 1 a). Bewegung: langsam, torkelnd, wühlt
in Algen- und Bakterienhäufchen. Nahrung: Algen. Manchmal häufig
in Weidetümpeln.
Abb. la-d. Platyophrya
citrina nach Lebendbeobachtungen. Infraciliatur und Silberliniensystem nach trockener Silberimprägnation. 1 a — Ansicht von links lateral.
Skala 20 (.im. 1 b, c — Körperquerschnitt proximal und subterminal. 1 d — I n f r a ciliatur und Silberliniensystem der rechten Seite
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W. F O I S S N E R
Somatische und orale Infraciliatur und Silberliniensystem (Abb. 1 d)
gleichen weitgehend Platyophrya vorax (s. die bei F o i s s n e r 1978
in Abb. 3 noch als Platyophrya sp. bezeichnete P. citrina).
P. citrina bildet bereits wenige Stunden nach dem Fang leicht orange
gefärbte, kugelförmige Cysten, die von einer sich mit MP intensiv blau
färbenden Ectocyste und einer sich nicht anfärbenden Entocyste umgeben sind. Bei der Konjugation vereinigen sich die Partner mit dem
Mund.
D i s k u s s i o n : P. citrina steht P. lata Kahl, 1930-35 nahe, deren
Dorsallinie eine ähnliche Form aufweist. Sie unterscheidet sich von
dieser durch die Färbung, die Anzahl der Kineten, den Körperquerschnitt und die Größe.
Pt
cv
Abb. 2 a-b. Platyophrya
dubia nach
Lebendbeobachtungen.
Infraciliatur
nach
trockener
Silberimprägnation.
2 a — Ansicht von rechts lateral. Skala 14 um. 2 b — Dorsalansicht
Platyophrya
dubia nov. spec. (Abb. 2 a, b)
D i a g n o s e : 25-32 jim große, sehr flexible, rechts mäßig gewölbte,
links fast ebene Platyophrya mit etwa 20 Zoochlorellen im hyalinen
Entoplasma. Gestalt spindelförmig. Ventral- und Dorsallinie mäßig konvex. Rechts 7-9 dicht, links 6-7 locker bewimperte Somakineten. 4-5
adorale Membranellen.
L o c u s t y p i c u s : Häufig in einem stark eutrophen Kleingewässer (Tümpel 52 a) östlich des Wallackhauses (Großglockner-Hochalpenstraße, etwa 2300 m ü.d.M.).
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CILIATEN DES
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33
M o r p h o l o g i e : Mundeingang spaltförmig, fast apikal gelegen,
von kurzen, in vivo schwierig erkennbaren Trichiten umgeben. Während
der Nahrungsaufnahme kann er sich auf etwa das Doppelte seines
normalen Durchmessers erweitern. Makronucleus kugelförmig, mit undeutlichen, länglichen Nucleolen, zentral gelegen. Mikronucleus ellipsoid, stark lichtbrechend, dem Makronucleus dicht anliegend. Pellicula
durch die leicht rechtsspiralig verlaufenden Somakineten stark gekerbt.
Cilien etwa 6 jxm lang, am distalen Pol ein etwa 5 |im durchmessendes
wimperloses Feld. Zwischen den Kineten zylindroide Protrichocysten.
Kontraktile Vakuole leicht subterminal. Entoplasma farblos, mit fettig
glänzenden, kugelförmigen Einschlüssen. Nur wenige Nahrungsvakuolen
mit Algenresten. Meist einige unregelmäßig geformte Defäkationsvakuolen (?) mit orange gefärbten Einschlüssen (Abb. 2 a).
Die somatische und orale Infraciliatur sowie das Silberliniensystem
gleichen weitgehend dem von P. citrina (s. oben).
D i s k u s s i o n : Diese Species unterscheidet sich von P. vorax
(s. unten) durch die geringere Größe, den stärker abgeflachten, spindelförmigen Körper, den mehr apikal gelegenen Mund sowie durch den
Besitz von Zoochlorellen. Von P. nana Kahl, 1926 unterscheidet sie sich
durch die Körperform, die kräftigere Kerbung der Pellicula und durch
die Zoochlorellen. Da aber diese Merkmale bei Platyophrya sehr variabel sind, ist nicht auszuschließen, daß P. dubia nur ein ökotyp einer
der oben erwähnten Arten ist, wenn man die Zoochlorellen nicht als
Specieskriterium gelten läßt. Von Woodruffia viridis Gelei, 1954, die
ebenfalls Zoochlorellen besitzt, ist P. dubia klar zu trennen, da W. viridis bedeutend größer ist, über 30 Somakineten und etwa 10 adorale
Membranellen besitzt (vgl. G r o 1 i è r e 1975).
Platyophrya
hyalina nov. spec. (Abb. 3 a-c)
D i a g n o s e : 36-45 jxm große, sich nach distal verschmälernde,
rechts konvexe und links konkave Platyophrya mit bisquittförmigem
Körperquerschnitt und stark gekerbter Pellicula. Oralapparat auf die
Ventralseite verlagert, mit 4-5 adoralen Membranellen. Makro- und
Mikronucleus von einer deutlich erkennbaren Membran umgeben. Zwischen Chromatin und Membran körnchenartige, anfärbbare Strukturen
(Bakterien?). Rechts 7-9 dicht, links 6-7 locker bewimperte Somakineten.
L o c u s t y p i c u s : Vereinzelt zwischen Moosen eines reinen Bächleins südlich des Heldendenkmales (Probenahmeort 26) (Großglockner-Hochalpenstraße, etwa 2300 m ü.d.M.).
M o r p h o l o g i e : P. hyalina ist auf der Höhe des weit nach ventral
versetzten, merkbar eingesenkten, von zarten Trichiten gestützten Oralapparates am breitesten und verschmälert sich nach distal allmählich.
î — A c t a P r o t o z o o l . 1/80
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34
W. F O I S S N E R
Das sich mit Orcein intensiv färbende Chromatin des Makronucleus
wird in weitem Abstand von einer Membran umgeben, die auch den
ellipsoiden Mikronucleus einschließt. Das ist typisch für Platyophrya
( F o i s s n e r 1978), aber nicht bei allen Arten so klar zu sehen. Pellicula durch die leicht rechtsspiralig verlaufenden Somakineten stark ge-
Pt
3c
Abb. 3 a-c. Platyophrya
hyalina
nach Lebendbeobachtungen. Infraciliatur nach trockener Silberimprägnation.
3 a — Ansicht von rechts lateral. Skala
20 |xm. 3 b — Ventralansicht. Skala
20 ixm. 3 c — Querschnitt in Körpermitte
kerbt. Zwischen den Wimperreihen Protrichocysten. Kontraktile Vakuole deutlich subterminal, Exkretionsporus auf der rechten Seite, etwa
4 um vom distalen Ende entfernt. Entoplasma meist mit vielen Nahrungsvakuolen mit Algen und farblosen Granula. Kriecht unruhig auf
Detritushäufchen und am Oberflächenhäutchen des Sammelglases
umher.
Somatische und orale Infraciliatur (Abb. 3 b) und Silberliniensystem
ähnlich wie bei P. vorax (s. F o i s s n e r 1978). In Folge der starken
Verlagerung des Mundes stoßen aber nur 3 Kineten an den rechten
Mundrand, während es bei den anderen hier beschriebenen Arten stets
4-5 sind.
D i s k u s s i o n : Unterscheidet sich hinsichtlich der Körperform von
allen bisher bekannten Vertretern der Gattung. Die Stellung des Mundes, die starke Kerbung der Pellicula und die Größe ähneln P. angusta
Kahl, 1926, bei der nach B u i t k a m p (1977) die adoralen Membranellen fehlen sollen.
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CILIATEN DES
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35
procera nov. spec. (Abb. 4 a-c)
D i a g n o s e : 45-81 fim große, lang ovale, etwa 2 : 1 abgeflachte,
leicht kontraktile und metabolische Platyophrya, deren Pellicula durch
die leicht rechtsspiralig verlaufenden Somakineten deutlich gekerbt
wird. Dorsallinie konvex, Ventrallinie leicht konkav. Rechts 7-9 dicht,
links 6-7 locker bewimperte Somakineten. Oralapparat auf die Ventralseite verschoben, mit 4-6 adoralen Membranellen.
Abb. 4 a-c. Platyophrya
procera nach
Lebendbeobachtungen. Infraciliatur und
Silberliniensystem nach trockener Silberimprägnation. 4 a — Ansicht von
rechts lateral. Skala 30 um. 4 b —
Infraciliatur und Silberliniensystem der
linken Seite. 4c — Körperquerschnitt
L o c u s t y p i c u s : Vereinzelt zwischen Moosen eines reinen Bächleins am Weg in das Rotmoos (Probenahmeort 4) (Großglockner-Hochalpenstraße, etwa 1200 m ü.d.M.).
M o r p h o l o g i e : Oralapparat von schwierig erkennbaren, kurzen
Trichiten umgeben. Makronucleus 8-10 [im im Durchmesser, mit vielen
kleinen Nucleolen und dicht anliegendem, spindelförmigem Mikronucleus.
Cilien der Dikineten lang und weich. Dicht unter der Pellicula rundliche Protrichocysten. Kontraktile Vakuole und Exkretionsporus leicht
subterminal. Entoplasma sehr hyalin, enthält neben wenigen Nahrungsvakuolen mit Algenresten meist einige größere orange gefärbte Einschlüsse und mäßig viele farblose, glänzende Granula. Gleitet langsam
auf Pflanzenresten und am Oberflächenhäutchen des Sammelglases. Somatische und orale Infraciliatur und Silberliniensystem sehr ähnlich wie
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36
W. F O I S S N E R
bei P. vorax (s. F o i s s n e r 1978). Basalkörper paarig angeordnet, auf
der locker bewimperten linken Seite trägt aber nur ein Basalkörper
eines Paares ein Cilium. Das trifft vielleicht auch auf die anderen hier
beschriebenen Arten zu!
D i s k u s s i o n : P. procera unterscheidet sich von den anderen
Arten der Gattung, deren Mund auf die Ventralseite verlagert ist, durch
die ungewöhnlich schlanke Körperform und den spindelförmigen Mikronucleus.
Platyophrya vorax Kahl, 1926 (Abb. 5 a, b, s. auch Abb. 1, 2 in
F o i s s n e r 1978)
M o r p h o l o g i e u n d D i s k u s s i o n : Ergänzend zur Beschreibung der Infraciliatur und des Silberliniensystems ( F o i s s n e r 1978)
hier noch Angaben zur Morphologie der lebenden Tiere. Größe
50-65 )j.m. Körper weich, verformbar. Gestalt stark vom Ernährungszustand abhängig. Trophont breit sackartig, kaum abgeflacht, Pellicula
glatt (Abb. 5 b). Theront lang oval, linke Körperseite manchmal deutlich
Abb. 5 a-b. Platyophrya vorax nach Lebendbeobachtungen. Infraciliatur nach trokkener Silberimprägnation. 5 a — Theront rechts lateral. Skala 25 um. 5 b — Trophont links lateral. Skala 30 um
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CILIATEN DES
37
GROSSGLOCKNERGEBIETES
abgeflacht, Pellicula durch die Wimperreihen kräftig gekerbt (Abb. 5 a).
Mund leicht subpolar, von mäßig deutlichen Trichiten umstellt, kann
bei der Nahrungsaufnahme stark erweitert werden. Makronucleus kugelförmig, fein granuliert, mit deutlicher, vom Chromatin leicht abgesetzter Membran, zentral bis subzentral gelagert. Mikronucleus spindelförmig, auffallend stark lichtbrechend, dem Makronucleus dicht anliegend. Zwischen den Wimperreihen rundliche Protrichocysten. Kontraktile Vakuole terminal, von vielen glänzenden Granula umgeben.
Entoplasma farblos, stets mit vielen schollenartigen, farblosen Granula
und einigen farblosen und orangeroten, fettig glänzenden, kugelförmigen Einschlüssen. Stark ernährte Individuen oft leuchtend grün durch
gefressene Algen. Nach W e n z e l (1953) kommen auch Stämme mit
Zoochlorellen vor. Bewegung rasch, mit dem proximalen Körperteil werden bohrende Bewegungen in Detritushäufchen durchgeführt.
2. Familie Colpodidae
Ehrenberg
Tabelle 1
Biometrische Charakteristik von Colpoda ovinucleata und Colpoda variabilis. Kinetenzahl nach
trocken versilberten Individuen. Alle anderen Werte nach naß imprägnierten Exemplaren
Parameter
Länge
Breite praeoral
Länge vom proximalen Pol bis zum
Beginn des Oralapparates
Länge vom proximalen Pol bis zum
Ende des Oralapparates
Makronucleus
Mikronucleus
Länge der linken
Polykinete
Durchmesser des Exkretionsporus
Gesamtanzahl der Kineten
Anzahl der postoralen Kineten
Extremwerte
(f*m)
Mittelwerte
((xm)
80-133
72-91
20-53
38-61
28-40
17-31
38-53
31-42
24-38x13-20
15-20x15-20
4.0-4.2x2.7-3.0
2.0-3.0x1.5-2.7
20.0-22.6
17.0-22.6
3.0
2.8-4.0
80-85
50-58
19-26
12-15
100.2
76.9
42.9
44.4
33.0
23.0
47.0
37.1
31.2X16.1
18.1x16.7
4.05x2.8
2.6x2.2
21.2
19.2
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-
3.4
81.9
54.0
22.5
14.4
Anzahl der
untersuchten
Individuen
11
16
9
11
6
11
6
9
6
8
4
7
4
10
1
5
8
11
9
10
38
W. F O I S S N E R
Colpoda ovinucleata
nov. spec. (Abb. 6 a-g, 10-16, 18, Tab. 1).
D i a g n o s e : In vivo 85-155 ^im große, etwa 35 fxm dicke, nierenförmige, rechts fast ebene, links deutlich gewölbte Colpoda, deren
Dorsalsack distal noch eine kleinere sackartige Vorwölbung besitzt.
Oralapparat mit großem, in der proximalen Körperhälfte gelegenem,
halbmondförmigem Vestibulum und gerade zur Körperlängsachse orientierter linker Polykinete. Etwa 8 bewimperte Kineten an der Innenwand
des Vestibulums. Makro- und Mikronucleus ellipsoid (etwa 2 : 1 ) . Durchschnittlich 60 Somakineten und 22 postorale Kineten, die in der tiefen
Diagonalrinne bis zum Exkretionsporus ziehen. Entoplasma mit plattenförmigen, glänzenden Kristallen und schwärzlichen Granula. Alle Cilien
paarig.
L o c u s t y p i c u s : Häufig in stark eutrophen Kleingewässern
(Tümpel 67) am Weg zur Pfandlscharte (Großglockner-Hochalpenstraße,
etwa 2200 m. ü.d.M.).
M o r p h o l o g i e : Körperform variabel, meist deutlich nierenförmig, praeoral schmäler als postoral (Abb. 6 a, 10). Am Eingang zum
Oralapparat, von dem eine hohlkehlenartige Rinne diagonal über die
linke Seite bis zum distalen Pol zieht, tief eingebuchtet. Die distale
Körperhälfte erscheint dadurch sackartig aufgewölbt. Am unteren Ende
dieses Sackes wölbt sich noch ein kleinerer, sehr leicht deformierbarer
Sack vor, in dem die kontraktile Vakuole liegt (Abb. 6 a, d, 10, 13, 14).
Dieser Sack war in den Präparaten stets faltig deformiert und verschwindet auch in vivo nach Auflegen des Deckglases sehr schnell. Rechte
Polykinete des Oralapparates aus unbekannten Gründen kaum imprägnierbar. Distale Hälfte des Vestibulumeinganges aufgewölbt, wodurch exakt dieselben Mundverhältnisse wie bei Tillina canalifera entstehen (s. T u r n e r 1937). Makronucleus rechts des Vestibulums gelegen,
mit netzartig angeordnetem Chromatin. Mikronucleus dem Makronucleus genähert und in nassen Silberpräparaten von einer auffällig
dicken Membran umgeben (Abb. 6 a). Pellicula durch die Wimperreihen deutlich gekerbt. Sie ist zwischen je 4 Dikineten bläschenartig
aufgewölbt. Darunter liegen die ca. 3 fim langen Protrichocysten, die
nach dem Ausstoß eine dünne Hülle um das Tier bilden. Die kontraktile
Vakuole entsteht durch Zusammenfließen vieler kleinerer Vakuolen.
Exkretionsporus leicht subterminal; Cytopyge etwas oberhalb der kontraktilen Vakuole. Im Entoplasma Tausende, 3-8 um große, gelbgrün
glänzende, plattenförmige Kristalle verschiedener Form und Größe.
Außerdem 1-3 jxm große, unregelmäßig sphaerische, schwärzliche Granula, die besonders im Dorsalsack gehäuft auftreten. Die etwa 10 fxm
großen Nahrungsvakuolen enthielten Bakterien und Phytoflagellaten.
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Abb. 6 a-g. Colpoda ovinucleata. 6 a — Ansicht von rechts lateral nach Lebendbeobachtungen. Skala 50 (.im. 6 b — Infraciliatur ventral-rechts lateral. Nasse Silberimprägnation. 6 c — I n f r a c i l i a t u r links lateral. Nasse Silberimprägnation.
6 d — Schematisierter Längsschnitt nach Lebendbeobachtungen. 6 e, f — Teile der
I n f r a c i l i a t u r und des Silberliniensystems der rechten Seite ohne und mit P r o t r i chocysten. Trockene Silberimprägnation. 6 g
— Teil der Infraciliatur und
des Silberliniensystems im Bereich der Diagonalrinne. Trockene Silberimprägnation
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40
W. F O I S S N E R
Bewegung im freien Wasser rasch, unter Rotation um die Längsachse.
An Detritushäufchen oft stillstehend oder kriechend und Nahrung
einstrudelnd.
Die Infraciliatur (Abb. 6 b, c, 10-12, 14-16) gleicht mit Ausnahme
der biometrischen Werte (Tab. 1) sehr jener von Tillina
magna
(s. L y n n 1976) bzw. Colpoda spiralis (s. N o v o t n y et al. 1977).
Von den durchschnittlich 22 postoralen Kineten verlaufen etwa 15 in
der Diagonalrinne, die sich distal immer mehr ausflacht. Dieses postorale Kinetenband ist bis zum Exkretionsporus erkennbar, da die Basalkörper sehr eng stehen. Im distalen Polbereich sind die Basalkörperpaare unregelmäßig angeordnet.
Das Silberliniensystem (Abb. 6 e-g, 15, 18) entspricht dem Typus
der Gattung (s. K l e i n 1929, F o i s s n e r 1978). Das Gitter ist meist
sehr unregelmäßig, was in Zusammenhang mit der Protrichocystenregeneration stehen mag. Die Relationskörper dieser Organellen liegen
in den Silberlinien (Abb. 6 f). Im distalen Polbereich und bei der
praeoralen Nahtlinie ist das Silberliniensystem engmaschig und nicht
orientiert. In der Diagonalrinne bildet es ziemlich regelmäßige, orthogonale Maschen (Abb. 6 g).
D i s k u s s i o n : Faßt man die Genera Colpoda und Tillina im Sinne
von G r u b e r (1880) und K a h l (1930-35) auf, so ist meine Art in
das Genus Colpoda zu stellen. T u r n e r (1937), G e l e i (1954) und
L y n n (1976) stellten zu Tillina allerdings Arten, die statt eines
röhrenförmigen Schlundes, wie er nach G r u b e r (1880), B r e s s l a u
(in K a h l 1930-35) und D i n g f e l d e r (1962) für Tillina typisch ist,
ein halbmondförmiges Vestibulum besitzen. Dadurch wird die Trennung
dieser Genera unscharf, da auch die Radiärkanäle der kontraktilen
Vakuole, die G r u b e r (1880) bei seiner Art vermutlich übersehen
hat, nicht bei allen Species vorhanden sind ( D r a g e s c o 1972). Das
halbmondförmige Vestibulum der T. magna von G e l e i (1954) veranlaßte bereits D i n g f e l d e r (1962), sie als Subspecies primitiva abzutrennen. Mir erscheint es aber richtiger, die Arten von G e l e i (1954)
und L y n n (1976) mit T. canalifera zu identifizieren, da T u r n e r
(1937) bei dieser Species das typische halbmondförmige Vestibulum als
Erster klar dargestellt hat. Es bleibt aber fraglich, ob man diese Art
nicht zu Colpoda oder Bresslaua stellen sollte. Auch C. ovinucleata
könnte man zu Bresslaua stellen, da K a h l (1930-35) eine Species
(Bresslaua sp.) in dieses Genus einordnete, bei der das Vestibulum nur
etwa die Hälfte des praeoralen Abschnittes einnimmt.
Innerhalb der Gattung Colpoda weist die neue Art nur zu C. reniformis
Kahl, 1930-35 nähere Verwandtschaft auf. Von dieser
90-100 (im großen Moosform unterscheidet sie sich durch den Dorsal-
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CILIATEN DES
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41
sack, das etwas größere Vestibulum, den mehr ellipsoiden Makronucleus,
die weniger deutlichen Protrichocysten, das ungefärbte Entoplasma,
die geringere Abflachung und die Entoplasmaeinschlüsse. Die Ubereinstimmungen sind — wenn man eine gewisse Subjektivität der Beobachter miteinrechnet — allerdings beträchtlich. Sollte sich bei der
Moosform von K a h l ein ähnlicher Dorsalsack finden, könnte man
C. ovinucleata vielleicht mit dieser synonymisieren.
Colpoda rotunda nov. spec. (Abb. 7 a-d)
D i a g n o s e : 130-170 fim große, sehr breit ovale, wenig abgeflachte
und beim Oralapparat kaum eingebuchtete Colpoda mit kugelförmigem
Makronucleus, dessen Nucleolen netzförmig angeordnet sind. Ein kugelförmiger Mikronucleus. Etwa 45 Kineten.
L o c u s t y p i c u s : Vereinzelt in einer Lithotekne (Tümpel 38)
zwischen Knappenstube und Hochtor (Großglockner-Hochalpenstraße,
etwa 2400 m ü.d.M.).
M o r p h o l o g i e : Stark ernährte Individuen fast kugelförmig. Am
Eingang zum Vestibulum nur wenig, manchmal überhaupt nicht eingebuchtet. Oralapparat verhältnismäßig klein, gattungstypisch aufgebaut, stets vor der Körpermitte liegend. Pellicula glänzend, da zwischen
je 2 Kineten 2-3 Reihen zylindroider Protrichocysten liegen (Abb. 7 d).
Kontraktile Vakuole terminal, rechts der Medianen, von vielen leicht
rötlichen Granula umgeben. Entoplasma dicht mit Nahrungsvakuolen
mit Bakterien und unregelmäßig kugelförmigen, kompakten Einschlüssen gefüllt. Bewegung auffallend langsam, manchmal auf der
Stelle kreisend. Vermehrung in Teilungscysten.
Wegen des seltenen Vorkommens konnte die Infraciliatur nicht genau studiert werden. Es sind ca. 45 Kineten vorhanden. Etwa 18 stoßen
am Kiel zusammen, ca. 8 ziehen vom Oralapparat weg. Sie sind von
den Somakineten der linken Seite nur undeutlich abgesetzt, da keine
Diagonalrinne vorhanden ist. Silberliniensystem (Abb. 7 c) gattungstypisch ausgebildet (vgl. F o i s s n e r 1978).
D i s k u s s i o n : Diese Species steht Colpoda cucullus nahe. Sie unterscheidet sich von dieser durch die Größe, die Struktur des Makronucleus, die geringere Einziehung beim Oralapparat, die größere Anzahl
der Wimperreihen und die geringere Abflachung. Keines dieser Merkmale reicht für sich allein aus, um die Aufstellung einer neuen Art zu
rechtfertigen, da C. cucullus sehr variabel ist (vgl. die Angaben von
R o u x 1901, K l e i n 1929, K a h l 1930-35, B u r t 1940, T u f f r a u
1952, W e n z e l 1953, G e l i e r t 1955, V ö r ö s v a r y 1950, R e u t e r
1961, H a s h i m o t o 1966).
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W. F O I S S N E R
Abb. 7 a-d. Colpoda rotunda nach Lebendbeobachtungen (a, d) und trockener
Silberimprägnation (b, c). 7 a — Ansicht von rechts lateral. Skala 75 jxm. 7 b —
Infraciliatur der Dorsalseite. 7 c — Teil der Infraciliatur und des Silberliniensystems. 7 d — Teil der Pellicula mit Protrichocysten
Colpoda variabilis nov. spec. (Abb. 8 a-g, 17, 19-24, Tab. 1)
D i a g n o s e : In vivo 80-120 (im große, etwa 30 fxm dicke, plump
nierenförmige, rechts fast ebene, links deutlich gewölbte Colpoda, deren
Dorsalsack durch die tief eingesenkte, bis zum distalen Körperpol reichende Diagonalrinne scharf akzentuiert ist. Oralapparat mit geräumigem, in der proximalen Körperhälfte gelegenem, trichterförmigem
Vestibulum und leicht schräg zur Körperlängsachse orientierter linker
Polykinete. Etwa 8 bewimperte Kineten an der rechten Innenwand des
Vestibulums. Makronucleus kugelförmig, Mikronucleus ellipsoid. Durchschnittlich 40 Somakineten und 14 postorale, sehr eng nebeneinander
verlaufende Wimperreihen. Alle Basalkörper sind paarig angeordnet.
Protrichocysten sehr auffällig, verquellen nicht zu einer strukturlosen
Hülle. Entoplasma mit vielen farblosen Granula. Ernährt sich von
Bakterien und Phytoflagellaten.
L o c u s t y p i c u s : Häufig in einem stark eutrophen Kleingewässer
(Tümpel 1) östlich des Wallackhauses (Großglockner-Hochalpenstraße,
etwa 2200 m ü.d.M.).
M o r p h o l o g i e : Körperform sehr variabel (Abb. 8 a-c), meist
plump nierenförmig, praeoral nur wenig schmäler als postoral, machmal
praeoral weit vorspringend (Abb. 8 a). Am Eingang zum Oralapparat
tief eingebuchtet. Von ihm zieht eine tiefe Rinne, die bei stark ernähr-
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Abb. 8 a-g. Colpoda variabilis. 8 a — Ansicht von rechts lateral. Nach Lebendbeobachtungen. Skala 50 jxm. 8 b — Infraciliatur ventral-rechts lateral. Nasse Silberimprägnation. 8 c — Infraciliatur links lateral. Nasse Silberimprägnation, Körperform nach Lebendbeobachtungen. 8 d — Schematisierter Längsschnitt nach
Lebendbeobachtungen. 8 e — Teil der Pellicula mit Protrichocysten. 8 f — ausgestoßene Protrichocysten nach MP-Färbung. 8 g — Individuum mit Protrichocystenhülle
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44
ten Tieren allerdings fast verschwindet, diagonal über die linke Körperseite bis zum distalen Pol. Rechte Polykinete des Oralapparates aus
unbekannten Gründen kaum imprägnierbar. Die distale Hälfte des
Vestibulumeinganges ist so wie bei C. ovinucleata aufgewölbt. Makronucleus kugelförmig bis leicht ellipsoid, rechts des Vestibulums gelegen,
mit großen runden und länglichen Nucleolen. Pellicula zwischen je
4 Basalkörperpaaren bläschenartig gewölbt. Protrichocysten etwa 3 |im
lang, 1-3 Reihen zwischen je 2 Wimperreihen (Abb. 8 e). In vivo sehr
auffallend und mit MP stark rot anfärbbar. Nach dem Ausstoß 4-6 [im
lang, in Opalblaupräparaten auch bis 20 (im. Form verschieden, Stäbchen- bis kommaförmig nach MP-Färbung und Silbernitratimprägnation
(Abb. 8 f), fadenförmig in Opalblaupräparaten. Kontraktile Vakuole
und Exkretionsporus leicht subterminal. Im freien Wasser ein gewandter, mäßig rascher Schwimmer. An Detritushäufchen oft stillstehend
oder kriechend und Nahrung einstrudelnd.
Die Infraciliatur (Abb. 8 b, c, 17, 19-22) und das Silberliniensystem
(Abb. 17, 23, 24) gleichen mit Ausnahme der biometrischen Werte (Tab.
1) im wesentlichen C. ovinucleata (s. dort). Als wichtiger Unterschied ist
festzuhalten, daß die Somakineten vom Kiel links und rechts gleich
schräg wegziehen (Abb. 8 b, 17, 19, 20), während sie bei C. ovinucleata rechts steiler an ihn stoßen als links (Abb. 6 b, 10, 15).
D i s k u s s i o n : Diese Species steht wegen ihrer variablen Körperform 3 Arten nahe, nämlich C. praestans Penard, 1922, C. simulans
Kahl, 1930-35 und C. henneguyi Fabre-Domergue, 1888. Von C. praestans unterscheidet sie sich durch die Größe, den kaum nach
dorsal zurückweichenden Kiel, die regellos verteilten Nucleolen, die
mehr distal gelegene kontraktile Vakuole und den Biotop. Das Vestibulum ist von P e n a r d (1922) nicht klar dargestellt worden, scheint
aber beträchtlich kleiner zu sein. C. praestans in der Beschreibung von
K a h l (1930-35) könnte mit C. variabilis identisch sein, wenn sie nicht
einen deutlich ellipsoiden Makronucleus hätte. Colpoda simulans weicht
durch die Größe, den schräg zurückweichenden Kiel, die eigenartigen
Nucleolen, die weniger auffälligen Protrichocysten, das Fehlen eines
ausgeprägten Dorsalsackes und durch den Biotop von C. variabilis ab.
C. henneguyi ist nur 31-65 (im groß und distal am rechten Rand stark
eingebuchtet.
3. Familie Marynidae Poche
Maryna ovata Gelei, 1950 (Abb. 9 a-c, 25-30)
M o r p h o l o g i e : Den sorgfältigen Lebendbeobachtungen von G el e i (1950, 1954) und D i n g f e l d e r (1962) habe ich nichts hinzuzufügen. Die starke Formvariabilität (Fig. 9 a-c) kann ich bestätigen. Die
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K
Abb. 9 a-c. Maryna ovata nach Lebendbeobachtungen
(Körperform)
und trockener
Silberimprägnation
(Infraciliatur,
Silberliniensystem).
9 a — Ventralansicht. 9 b — Ansicht von rechts lateral. 9 c — Dorsalansicht
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46
W. FOISSNER
Infraciliatur und das Silberliniensystem sind vom G e l e i (1950) nur in
den Ansätzen richtig dargestellt worden. Die Infraciliatur läßt sich in
3 Abschnitte gliedern (Abb. 9 a-c, 25-29):
(1) Etwa 50 ( G e l e i gibt nur bis 36) praeorale Kineten stoßen entlang
der ventralen, leicht sigmoiden Nahtlinie und des apikalen Poles zusammen. Die Kineten der linken Seite stoßen spitzwinkelig an den Kiel,
die der rechten Seite verlaufen zuerst fast parallel mit der Nahtlinie.
Etwa 4 Kineten werden durch den Mund abgeschnitten. Die folgenden 7-10 Kineten ziehen rechts des Mundes vorbei und biegen postoral
leicht nach links. 4-5 dieser Kineten stoßen an die postorale Nahtlinie,
während die anderen erst am Rand der distalen Polfläche enden. Einige
Basalkörper dieser und der an sie seitlich anstoßenden mycteralen Kineten wandern auf die distale Polfläche hinaus. Die linke Abbiegung,
die auch von einem Zusammenrücken der Kineten begleitet ist, bewirkt
den linksspiraligen Verlauf der Wimperreihen. Die Cilien sind am
ganzen Körper paarig angeordnet und stehen im distalen Drittel etwa
doppelt so eng wie im proximalen Abschnitt.
(2) Links des Mundes beginnen die sehr dicht bewimperten mycteralen Kineten. Es sind ca. 15 schräg
45°) zur Körperlängsachse orientierte Wimperreihen. Die letzten 2 Reihen sind verkürzt und stoßen an
die durchlaufenden Kineten rechts des Oralapparates.
(3) An die mycteralen Kineten stoßen distal 25-30 Kineten zu je
etwa 6 Basalkörperpaaren. Die distalen Dikineten jeder Reihe sind
leicht vergrößert und bestehen vielleicht aus 4 sehr eng nebeneinander
liegenden Basalkörpern. Von ihnen entspringen die stark verlängerten,
sehr empfindlichen Caudalcilien.
Das Silberliniensystem (Abb. 9 a-c, 25-29) ist dem Weitmaschentyp
zuzuordnen. Jedes Basalkörperapaar wird von einer Zirkularfibrille umgeben, von der 4 Silberlinien zu den benachbarten 4 Basalkörperpaaren
ziehen. Die dadurch entstehenden Maschen sind wegen des spiraligen
Kinetenverlaufes shombisch. Entlang der prae-und postoralen Nahtlinie
verläuft eine nicht unterbrochene Silberlinie, die mit den Silberlinien
der anstoßenden Somakineten kontaktet. Die mycteralen Kineten werden durch umgekehrt Z-förmige Silberlinien verbunden. Auf dem unbewimperten proximalen Pol breitet sich ein unregelmäßiges Engmaschengitter aus, das von den Silberlinien der Somakineten gebildet wird.
Die distale Polfläche ist bis auf die oben erwähnten Basalkörperpaare unbewimpert und von einem feinmaschigen Silberliniennetz bedeckt.
Im Zentrum dieser Polfläche liegt der Porus der kontraktilen Vakuole.
Der Rand des Vestibulums wird von einem Engmaschengitter bedeckt.
Die Oralstrukturen konnten nicht genau erkannt werden. Eine typische
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CILIATEN DES
GROSSGLOCKNERGEBIETES
47
Polykinete ist oft gut imprägniert, in vivo wurde auch eine zweite Polykinete festgestellt (vgl. G e 1 e i 1950).
D i s k u s s i o n : Meine Untersuchungen bestätigen die von G e l e i
(1954) und D i n g f e l d e r (1962) vorgenommene Einordnung der Marynidae in die Colpodida (vgl. F o i s s n e r 1978) und geben keine Hinweise dafür, daß diese Familie in die Trichostomatida einzuordnen wäre
( C o r l i s s 1977). Die von D i n g f e l d e r (1962) vorgeschlagene Ableitung der Marynidae von den Colpodidae kann ich voll unterstützen. Zu
den von ihm angeführten homologen Merkmalen sind auf Grund der
vorliegenden Untersuchungen folgende hinzuzufügen:
(1) Das Silberliniensystem entspricht dem Typus der Colpodidae
(vgl. F o i s s n e r 1978).
(2) Die praeorale Nahtlinie ist homolog dem Kiel der Colpodidae.
Die auffällige Länge dieser Nahtlinie zeigt, daß der Mund im Verlaufe
der Evolution nach distal gewandert sein muß, zusammen mit den Kineten der Diagonalrinne.
DANKSAGUNG
Mit dankenswerter finanzieller Unterstützung des MaB-6 Projektes der österreichischen Akademie der Wissenschaften.
SUMMARY
The morphology, infraciliature, and silverline system of some new or little
known Colpodida occurring in small water bodies of the Großglockner area (Hohe
Tauern, Austria) is described. The following species are new to science: Platyophrya citrina nov. spec., Platyophrya
dubia nov. spec., Platyophrya
hyalina nov.
spec., Platyophrya
procera nov. spec., Colpoda ovinucleata nov. spec., Colpoda rotunda nov. spec., and Colpoda variabilis nov. spec. The infraciliature and the silverline system of Maryna ovata demonstrate that the Marynidae are closely related
to the Colpodidae.
LITERATUR
B u i t k a m p U. 1977: Die Ciliatenfauna der Savanne von Lamto (Elfenbeinkuste).
Acta Protozool., 16, 249-276.
B u r t R. L. 1940: Specific analysis of the genus Colpoda with special reference
to the standardization of experimental material. Trans. Am. Microsc. S o c ,
59, 414-432.
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48
W. F O I S S N E R
C o r l i s s J. O. 1953: Silver impregnation of ciliated protozoa by the ChattonLwoff technic. Stain. Technol., 28, 97-100.
C o r l i s s J. O. 1977: Annotated assignment of families and genera to the orders
and classes currently comprising the corlissian scheme of higher classification for the phylum ciliophora. Trans. Am. Microsc. Soc., 96, 104-140.
D i n g f e l d e r J. H. 1962: Die Ciliaten vorübergehender Gewässer. Arch. Protistenk., 105, 509-658.
D r a g e s c o J. 1970: Ciliés libres du Cameroun. Ann. Fac. Sci. Yaounde (Hors
Série), pp. 1-141.
D r a g e s c o J. 1972: Ciliés libres de l'Ouganda. Ann. Fac. Sci. Cameroun, 9,
87-126.
F a b r e - D o m e r g u e P. L. 1888: Recherches anatomiques et physiologiques sur
les infusoires ciliés. Ann. Sci. nat., 5, 1-140.
F o i s s n e r W. 1976: Erfahrungen mit einer trockenen Silberimprägnationsmethode zur Darstellung argyrophiler Strukturen bei Protisten. Verh. Zool.Bot. Ges. Wien, 115, 68-79.
F o i s s n e r W. 1978: Das Silberliniensystem und die Infraciliatur der Gattungen
Platyophrya
Kahl, 1926, Cyrtolophosis
Stokes, 1885 und Colpoda O.F.M.,
1786: Ein Beitrag zur Systematik der Colpodida (Ciliata, Vestibulifera). Acta
Protozool., 17, 215-231.
F o i s s n e r W. 1979: Methylgrün-Pyronin: Seine Eignung zur supra vitalen Übersichtsfärbung von Protozoen, besonders ihrer Protrichocysten. Mikroskopie,
35, 108-115.
F o i s s n e r W. 1980: Artenbestand und Struktur der Ciliatenzönose in alpinen
Kleingewässern (Hohe Tauern, Österreich). Arch. Protistenk. (im Druck).
G e l e i J. v. 1950: Die Marynidae der Sodagewässer in der Nähe von Szeged.
XIV. Beitrag zur Ziliatenfauna Ungarns. Hidrol. Közl., 30, 107-119, 157-158.
G e 1 e i J. v. 1954: Über die Lebensgemeinschaft einiger temporärer Tümpel auf
einer Bergwiese im Börzsönygebirge (Oberungarn). III. Ciliaten. Acta biol.
Acad. Sci. hung., 5, 259-343.
G e l l é r t J. 1955: Die Ciliaten des sich unter der Flechte Paramelia
saxatilis
Mass. gebildeten Humus. Acta biol. Acad. Sci. hung., 6, 77-111.
G r o l i è r e C.-A. 1975: Contribution a l'etude des ciliés des sphaignes et des
etendues d'eau acides. I- Description de quelques especes de Gymnostomes,
Hypostomes, Hymenostomes et Heterotriches. Ext. Ann. Sta. Biol. Besse-enChandesse, 10, 265-297.
G r u b e r A. 1880: Neue Infusorien. Z. wiss. Zool., 33, 439-466.
H a s h i m o t o K. 1966: Stomatogenesis in resting cysts of Colpodidae. J. Protozool., 13, 383-390.
K a h l A. 1926: Neue und wenig bekannte Formen der holotrichen und heterotrichen Ciliaten. Arch. Protistenk., 55, 197-438.
K a h l A. 1930-35: Urtiere oder Protozoa. I. Wimpertiere oder Ciliata (Infusoria).
In: Die Tierwelt Deutschlands, (ed. Dahl F.) G. Fischer, Jena 886 pp.
K l e i n B. M. 1929: Weitere Beiträge zur Kenntnis des Silberliniensystems der Ciliaten. Arch. Protistenk., 65, 183-257.
L y n n D. H. 1976: Comparative ultrastructure and systematics of the Colpodida.
Fine structural specializations associated with large body size in Tillina
magna Gruber, 1880. Protistologica, 12, 629-648.
N o v o t n y R. T., L y n n D. H. und E v a n s F. R. 1977: Colpoda spiralis sp.
n., a colpodid ciliate found inhabiting treeholes (Colpodida, Ciliophora). J.
Protozool., 24, 364-369.
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CILIATEN DES GROSSGLOCKNERGEBIETES
49
P e n a r d E. 1922: Études sur les infusoires d'eau douce. Georg et Cie, Genève,
331 pp.
R e u t e r J. 1961: Einige faunistische und ökologische Beobachtungen über Felsentümpel-Ziliaten. Acta Zool. Fenn., 99, 1-42.
R o u x J. 1901: Faune infusorienne des eaux stagnantes des environs de Genève.
Kündig, Genève, 148 pp.
T u l f r a u M. 1952: La morphogenèse de division chez les Colpodidae. Bull. biol.
Fr. Belg., 86, 309-320.
T u r n e r J. P. 1937: Studies on the ciliate Tillina canalifera n. sp. Trans. Ara.
Microsc. Soc., 56, 447-456.
V ö r ö s v ä r y B. 1950: Die Ciliaten des "Kalânos"-Baches. Ann. Biol. Univ. szeged.,
1, 343-387.
W e n z e l F. 1953: Die Ciliaten der Moosrasen trockner Standorte. Arch. Protistenk., 99, 70-141.
4 — Acta Protozool. 1/80
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LEGENDEN ZU DEN TAFELN I-II
10-16, 18: Colpoda ovinucleata.
Infraciliatur und Silberliniensystem nach nasser
Silberimprägnation (10-12, 14, 16) und trockener Silberimprägnation (15, 18).
10, 11 — Ansicht eines Individuums von ventral-rechts lateral in verschiedener
Fokushöhe. 12 — Ventralansicht. 13 — freischwimmendes Exemplar, Phasenkontrast, Ansicht von links lateral. 14 — Seitenansicht. Der Pfeil weist auf die Diagonalrinne. 15 — Ventralansicht. 16 — Ansicht von links lateral. 18 — Teil der
Infraciliatur und des Silberliniensystems
17, 19-24: Colpoda variabilis. Infraciliatur und Silberliniensystem nach nasser Silberimprägnation (19, 20-23) und trockener Silberimprägnation (17, 24). 17 — Ansicht von ventral und proximal. 19 — Ansicht von ventral-rechts lateral. 20,
21 — Ansicht eines Tieres von ventral-rechts lateral in verschiedener Fokushöhe.
22 — Ansicht von links lateral. 23, 24 — Teile der Infraciliatur und des Silberliniensystems der rechten Seite. Die Pfeile weisen auf die Basalkörperpaare
25-30: Maryna ovata. Infraciliatur und Silberliniensystem nach trockener Silberimprägnation. 25 — Ventralansicht. Die Pfeile weisen auf die Ventralnaht. 26 —
proximaler Pol. Der Pfeil weist auf die Ventralnaht. 27 — Dorsalansicht. 28 —
distaler Körperteil. Der Pfeil weist auf eine Reihe vergrößerter Basalkörperapparate. 29 — Ansicht von rechts lateral. 30 — Lebendaufnahme im Phasenkontrast.
Das lange Gallertgehäuse ist gut erkennbar
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ACTA P R O T O Z O O L .
VOL.. 19, N o . 1
TAFEL I
W. Foissner
auctor phot.
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ACTA
P R O T O Z O O L . VOL.. 19, N o . 1
TAFEL I
auctor phot.
W. Foissner
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Protozoology Laboratory, Department of Zoology, University of Calcutta,
35 Ballygunge Circular Road, Calcutta 700019, India
A. K. C H A N D R A and A. C H O U D H U R Y
A New Opalinid Hegneriella mukundai sp. n. from an Old World
Hylid Kaloula pulchra taprobanica Parker from India
Received
21 May 1979
Synopsis.
Hegneriella
mukundai
sp. n. having body dimensions
93.65 jim ± 2.71 urn X 58.05 |im ± 2.24 ^m and nuclear dimensions
25.01 jim ± 1.74 nm X 17.72 |xm±0.49 nm with eight to ten nucleoli,
is described from Kaloula pulchra taprobanica Parker and also compared with those of H. dobelli Earl, 1971 and H. cheni Earl, 1972.
Since the erection of the genus Hegneriella by E a r l (1971), which
embraces only two species H. dobelli Earl, 1971 and H. cheni Earl,
1972, no other contribution to this genus has so far been made. The
present study records a third species of this genus, namely H. mukundai, sp. n., which is its first occurrence in the Old World.
In the course of this investigation, 517 host animals, Kaloula pulchra
taprobanica Parker were examined. Some harboured only flagellates,
others mixed populations of Zelleriella spp., Protoopalina spp. and
nyctotherans. Only four individuals had Hegneriella mukundai sp. n.
Neither Opalina spp. nor Cepedea spp. were encountered in this host.
Materials
and
Methods
The host specimens Kaloula pulchra taprobanica Parker were collected from
the district of Midnapore, West Bengal. The rectal contents of the hosts were
emptied into drops of 0.65% saline on slides. Living opalinids were examined
with the phase-contrast microscope. Part of the material was fixed in Schaudinn's
and stained with iron hematoxylin, after C h e n (1944). Silver impregnation after
C o r l i s s (1953) was used for the study of the infraciliature, even though iron
alum hematoxylin stain served well for this purpose. Measurements were made
on both living and stained opalinids, the composite drawing being made with
the aid of a camera lucida and microphotographs.
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52
A . K. C H A N D R A A N D A . C H O U D H U R Y
R e s u l t s
Typical trophozoites of Hegneriella mukundai sp. n. are oval; the
posterior section is broader than the anterior; ventral margin is slightly
concave towards the anterior side. The cell is uninucleate. The body
measures 81.7-107.5X47.3-73.1 jim (see Fig. 1).
The body is uniformly ciliated with short (2.5-5.1 }im) cilia. The ciliary
lines (14-19) run obliquely parallel to the anterioposterior axis, having interkinetal distances which average 1.7 jxm anteriorly and 3.4 near
the posterior pole.
The large, ovoid nucleus, ca. 25X18 fim, is central and placed obliquely to the longitudinal axis. Its variously-sized, round nucleoli are
scattered in the nucleoplasm, numbering eight to ten (Fig. 2).
Fig. 1. Hegneriella
mukundai
sp. n.
Fig. 2. Prolate nucleus of
Hegneriella
mukundai sp. n. showing nucleoli (arrows)
Discussion
Both Hegneriella dobelli and H. cheni were described from the same
anuran host: Bufo valliceps, but from Louisiana and Texas respectively. H. dobelli is broad and obliquely truncated at the anterior end,
the posterior end being comparatively narrower, whereas H. cheni has
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53
a broad not narrow posterior section. The nucleus of H. cheni is spindle-shaped, containing three nucleoli, while that of H. dobelli is large
and prolate, containing 10 nucleoli, which may reflect difference in the
number of chromosomes.
H. mukundai, inscribed to the memory of the reputed Indian protozoologist Prof. M. M. Chakravarty, has a single large nucleus, containing eight to ten nucleoli, and shows some resemblance to H. dobelli,
though different in dimensions (see Table 1). Measurements of the
three known species are compared in Table 1.
E a r l (1979) has commented, under the possible title of "unfinished
business", that H. cheni was bypassed by the collector, C h e n , enigmatically, since, in his opinion C h e n (1948) was extraordinarily competent and an experienced cytologist familiar with Zelleriella.
Table 1
A comparative mensural data of Hegneriella dobelli Earl, 1971, H. cheni Earl 1972 and
H. mukundai sp. n. in microns
Length of the body
Width of the body
Length of the nucleus
Breadth of the
nucleus
Range
Arithmetic
mean
Standard
deviation
Standard
error
H. dobelli
H. cheni
H. mukundai
62.0-133.0
86.0-168.0
81.7-107.5
98.2
142.0
93.6
23.10
2.3
2.2
2.7
24
H. dobelli
H. cheni
H. mukundai
50.0-75.0
44.0-89.0
47.3-73.1
61.0
70.7
58.0
14.00
1.4
0.7
2.2
23
H.
H.
H.
H.
H.
H.
30.0-42.0
27.0-50.0
17.0-37.4
13.0-21.0
12.3-24.0
13.6-18.7
34.1
36.6
25.0
16.2
16.9
17.7
5.70
6.77
4.20
1.92
0.6
0.7
1.7
0.4
0.3
0.4
0.80
0.2
dobelli
cheni
mukundai
dobelli
cheni
mukundai
—
10.52
—
8.68
—
—
—
11
—
14
6
—
27
4
—
10
21
Length of the cilia
H. mukundai
2.5-5.1
3.6
Distance between
the two ciliary lines
in the anterior region
H. mukundai
-
1.7
-
-
—
Dist nee between
the two ciliary lines
in the posterior
region
H. mukundai
-
3.4
-
-
-
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54
A. K. C H A N D R A A N D A. C H O U D H U R Y
ACKNOWLEDGEMENTS
Authors are greatly thankful to Dr. P. R. Earl for showing his great interest in this subject. Sincere thanks are also due to Professor Dr. Asok Ghosh,
Head of the Department of Zoology, University of Calcutta, for providing necessary laboratory facilities.
RÉSUMÉ
Hegneriella mukundai sp. n., avec les dimensions du corps: 93.65 ura ± 2.71 nm X
X 58.05 nm ± 2.24 fim, les dimensions du noyau: 25.01 pim ± 1.74 ura X 17.72 jmi ±
± 0.49 nm, et avec &-10 nucléoles, trouvée chez Kaloula pulchra taprobanica Parker, est décrite et comparée avec H. dobelli Earl, 1971 et H. cheni Earl, 1972.
REFERENCES
C h e n T. T. 1944: Staining nuclei and chromosomes in Protozoa. Stain Tech., 19,
83-90.
C h e n T. T. 1948: Chromosome in Opalinidae (Protozoa, Ciliata) with special reference to their behavior, morphology, individuality, diploidy, haploidy and
association with nucleoli. J. Morph., 83, 281-357.
C o r l i s s J. O. 1953: Silver impregnation of ciliated Protozoa by the ChaittonLwoff technic. Stain Tech., 28, 97-100.
E a r l P. R. 1971: Hegneriella dobelli gen. n., sp. n. (Opalinidae) from Bufo valliceps and some remarks on the systematic position of the opalinidae. Acta
Protozool., 9, 41-48.
E a r 1 P. R. 1972: Hegneriella cheni sp. n. and Opalina wessenbergi sp. n. (Protozoa).
Acta Protozool., 10, 267-268.
E a r 1 P. R. 1979: Personal communication.
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ACTA
VOL. 19 (No. 1)
PROTOZOOLOGICA
WARSZAWA, 30 III 1980
Laboratory of Protozoology, Department of Zoology, Marathwada University,
431 004, India
pp. 55-60
Aurangabad
P. A. L A L P O T U
Maupasella perionychis sp. n. a New Astomatous Ciliate from
the Earthworm Perionyx sansibaricus in India
Received
15 May 1979
Synopsis: A new astomatous ciliate Maupasella perionychis sp. n. (sub.
fam. Hoplitophryoidea
P u y t o r a c 1972) is described from the gut
in Maharashtra, India. It is
of an earthworm Perionyx sansibaricus
characterized by a long ribbon like body measuring 95.8 |xm X 18.9 ftm
on the average and with 26 rows of kineties on one side and 18-20 on
other. This is the first record of an astomatous ciliate from earth worm
of this region.
The genus Maupasella was erected by C£pede 1910 for a ciliate
which had been earlier recorded by Maupas from the alimentary canal
of an Algerian earthworm. Subsequently several forms have been described by different workers like K e i 1 i n (1920), B h a t i a and G u 1 a t i
(1927), H e i d e n r e i c h (1935 a, b). W i c h t e r m a n (1939), K a t a s h i m a (1952), P u y t o r a c (1952) and L o m (1959 a, b, c, 1961). P u y t o r a c (1972) in his review of the astomatous ciliates included this
genus in the subfamily Maupasellinae C6pede, 1910, family Maupasellidae Cheissin, 1930 under a new superfamily Hoplitophryoidea. He listed
16 species of this occasional change of direction or a progressive push
forward, rotating along their long axis.
The body of the ciliate is typically elongated, flat and ribbon shaped,
having more or less uniform width troughout (Fig. 1 1). The two ends
are rounded, the anterior one being more broadly rounded and the
posterior slightly narrower. At the anterior tip of the body is a typical spine-like outgrowth (about 3.5-4 (im long) which is characteristic and probably helps for attachment (Fig. 1 2). The body length is
about five times the width. The body is covered by a fairly thick pellicle which is quite flexible permitting body movements in all directions.
The cytoplasm appears granular and there is no apparent distinction
into ecto and endo plasm. The cytoplasm around the macronucleus is
vacuolar. The body is covered with a thick coat of cilia arranged in
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56
P . A. L A L P O T U
closely set longitudinal lines. The cilia are uniform all over the body
and are about 3-4 ^im in length on the average.
Fig. 1. Maupasella perionychis sp. n. 1 — Showing general morphology (based on Schauddin/Haematoxylin preparations), 2 — Showing
the spine-like structure at anterior end (based
on an organism stained with Lugol's solution
There are on an average 46 ciliary
rows, 26 of them on the ventral side (Pl.
I 3) and 20 on the dorsal side. The kineties converge at the anterior end on the
ventral side on a small triangular unciliated area, from where arises the spinelike adhesive organ (Pl. I 3, 4). The kineties run backwards more or less parallel
to one another and converge on a posterior suture coinciding with the posterior
end of the body.
The macronucleus is extremely long,
extending more than three-fourths of the
length of the body. It is ribbon shaped and
granular, with a smooth outline in most
cases (Fig. 1 1, 2). The micronucleus could
not be seen in any of the specimens genus from various annelids in different
parts of the world. Form India B h a t i a and G u 1 a t i (1972) were the only
authors todate who have recorded this genus. They found M. nova in
Pheretima posthuma and Pheretima
hawayana.
Material
and
Methods
The ciliates described were found in the gut of an earthworm Perionyx
sansibaricus during the course of a survey of the ciliate fauna of invertebrates of
Maharashtra, State, India. The organisms were examined in detail in living condition in temporary wax-sealed preparations. Observations in the living condition
were also made with the help of vital stains. Particularly neutral red and methylene blue were useful, besides Lugol's solution. Permanent preparations were
made with Heidenhain's iron haematoxylin and phospho-tungstic acid haematoxylin, following fixation in Schaudinn's fluid. Silver impregnation was done by
the dry method, after K l e i n (1958).
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N E W A S T O M A T O U S C I L I A T A MAUPASELLA
Description of Maupasella perionychis
(Plate I 3, 4, 5)
PERIONYCHIS
57
sp. n.
The ciliate was found in the gut of the earthworm, Perionyx sansibaricus collected from the viscinity of Aurangabad, Maharashtra. The
parasites were found mostly just above the middle of the body being
particularly concentrated in the post-clitellar region. The infection was
never heavy, even though the incidence was common.
In the living condition the ciliate moves about very slowly. The
movement is mostly spiral around the same point with an occasional
change of direction or a progressive push forward, rotating along their
long axis.
The body of the ciliate is typically elongated, flat and ribbon shaped,
having more or less uniform width throughout (Fig. 1 1). The two ends
are rounded, the anterior one being more broadly rounded and the posterior slightly narrower. At the anterior tip of the body is a typical
spine-like outgrowth (about 3.5-4 (im long) which is characteristic and
probably helps for attachment (Fig. 1 2). The body length is about five
times the width. The body is covered by a fairly thick pellicle which
is quite flexible permitting body movements in all directions. The cytoplasm appears granular and there is no apparent distinction into ecto
and endo plasm. The cytoplasm around the macronucleus is vacuolar.
The body is covered with a thick coat of cilia arranged in closely set longitudinal lines. The cilia are uniform all over the body and are about
3-4 fim in length on the average.
There are on an average 46 ciliary rows, 26 of them on the ventral
side (PI. I 3) and 20 on the dorsal side. The kineties converge at the
anterior end on the ventral side on a small triangular unciliated area, from
where arises the spine-like adhesive organ (PI. I 3, 4). The kineties run
backwards more or less parallel to one another and converge on a posterior suture coinciding with the posterior end of the body.
The macronucleus is extremely long, extending more than three-fourths
of the length of the body. It is ribbon shaped and granular, with
a smooth outline in most cases (PI. I 2). The micronucleus could not be
seen in any of the specimens examined as it is probably very small and
covered over by the micronucleus.
No contractile vacuoles were seen in any of the living or fixed
organisms.
The dimensions of the organism are shown in Table 1.
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58
P. A. L A L P O T U
Discussion
Of the various species listed by P u y t o r a c (1972) this species comes close to M. mucronata (C£pede, 1910 Puytorac, 1954 in the number
of the kineties i.e., about 46. However, the present species is much
longer in size than M. mucronata, measuring 66.2-132.3 ^m X 15.8-22.1
[Am (95.8 jim X 18.9 fim) as against 30-90 (im X 48 [j.m. Particularly, the
extremely elongated and narrow ribbon like body of the present species is characteristic with the length-breadth ratio being almost 5 : 1 .
The ciliate described above from Perionyx sansibaricus appears to
be similar to the elongated forms described by B h a t i a and G u 1 a t i
(1927) from two species of Pheretima, P. posthuma and P. hawayana
in its body dimensions and in the length of the macronucleus. These
authors have described that there are fine and closely set cilia disposed
in 27 longitudinal rows which could be "counted at the broader end of
Table 1
Dimensions of Maupasella perionychis sp. n. (all measurements in microns)
Particulars
Length of the body
Width of the body
Length of the macronucleus
Width of the macronucleus
Ratio of length of the body and
width of the body
Ratio of length of the body and
length of tne macronucleus
Minimum
Maximum
Average
66.2
15.8
53.6
3.0
132.3
22.1
110.3
4.8
95.8
18.9
80.3
3.5
5 1
1.2 1
the body". In the present organism there are about 26 rows on one side
and 18-20 on the other. In view of these and the fact that the number
and arrangement of the kineties is a major basis for taxonomic identity
of the various species at present, the present organism is considered
identical with the species described by B h a t i a and G u 1 a t i as M.
nova in spite of the fact that B h a t i a and G u 1 a t i described the
presence of many contractile vacuoles, while none are present in the
present organism.
C 6 p e d e (1910) gave the name M. nova for a parasite which had
earlier been discovered by Maupas from the alimentary canal of an
Algerian earthworm. Similar organisms were recorded later by several
workers, including B h a t i a and G u 1 a t i (1927). As M a c k i n n o n
and H a w e s (1961) stated "for many years thereafter apparently
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59
the same organism was remarked, always in oligochaetes, until a considerable number of distinct but related species had accumulated under
C é p è d e's name. Unfortunately, C é p è d e did not identify his hosts
and his description is defective in just those particulars by which the
species now recognized are distinguished, so that it is impossible to tell
which of these corresponds with that seen by the original author or,
in many cases, those seen by his successors. Thus the name that is so
common in the literature, M. nova, can no longer be applied to any species".
In view of the foregoing discussion it appears necessary not to apply
the name M. nova to any species and hence this ciliate is designated
as Maupasella perionychis sp. n. with M. nova sensu B h a t i a and
G u 1 a t i (1927) as its synonym.
Species: Maupasella perionychis sp. n.
Host: Perionyx
sansibaricus
Habitat: Alimentary canal
Locality: Aurangabad, Maharashtra, India.
The type slides are deposited in the Protozoology Laboratory, Department of Zoology, Marathwada University, Aurangabad 431 004, India.
ACKNOWLEDGEMENTS
The author is thankful to Dr. R. Krishnamurthy for his valuable guidance and
to Dr. R. Nagabhushanam for providing laboratory facilities. Thanks are also due
to the Marathwada University, Aurangabad for the award of U.G.C. Junior Research Fellowship.
RÉSUMÉ
Un nouveau Cilié astome Maupasella perionychis sp. n. (sous-famille Hoplitophryoidea Puytorac 1972) a été trouvé dans l'intestin du Lombric Perionyx
sansibaricus à Maharashtra, aux Indes. Son corps allongé en forme de ruban mesure
95.8 (xm X 18.9 jxm en moyenne et possède 26 rangs des cinétosomes d'un côté et
18-20 de l'autre. C'est la première découverte d'un Cilié astome chez un lombric
dans cette région.
REFERENCES
B h a t i a B. L. and G u l a t i A. N. 1927: On some parasitic ciliates from Indian
frogs, toads, earthworms and cockroaches. Arch. Protistenk., 57, 85-120.
C é p è d e C. 1910: Researches sur les Infusoires astomes. Anatomie, biologie,
éthologie, parasitaire, systématique. Arch. Zool. exp. gén., (Sér. 5) 3, 341609.
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60
P . A. L A L P O T U
C h e i s s i n E. 1930: Morphologische und systematische Studien über
Astomata
aus dem Baikalsee. Arch. Protistenk., 70, 531-618.
H e i d e n r e i c h E. 1935a: Untersuchungen an parasitischen Ciliaten aus Anneliden. Teil I: Systematik. Arch. Protistenk., 84, 315-392.
H e i d e n r e i c h E. 1935 b: Ergänzende Untersuchungen an parasitischen Ciliaten
aus Oligochaten. Arch. Protistenk., 528-532.
K a t a s h i m a R. 1952: Studies on the Astomata (Ciliata, Protozoa). II Zool. Mag.,
62,
22.
K e i 1 i n D. 1920: On the occurrence of a supplementary chromatic body in Mauppasella nova Cépède, an intestinal parasit of earthworms. Parasitology, 12.
K l e i n B. M. 1958: The "dry" silver method and its proper use. J. Protozool., 5,
99-103.
L o m J. 1959 a: Beiträge zur kenntnis der parasitischen ciliaten aus Evertebraten
III Neue-Arten der gattung Bütschliellopsis de Puytorac, 1954 und der gattung
Akidodes n. g. Arch. Protistenk., 103, 457-488.
L o m J. 1959 b: Beiträge zur kenntnis der parasitischen ciliaten aus Evertebraten
IV Neue Ciliaten aus der Familie Haptophryidae
Cépède, 1923, nebst einigen
Bemerkungen zum heutigen Stand dieser Gruppe. Arch. Protistenk., 104,
133-154.
L o m J. 1959 c: A contribution to the knowledge of astomatous ciliates. Mem.
Soc. Zool. Tchecosl., 23, 200-210.
L o m J. 1961: Some remarks on the morphology and taxonomy of astomatous ciliates from earthworms. Acta Soc. Zool. Bohem., 25, 167-180.
M a c k i n n o n D. L. and H a w e s R. S. J. 1961: An introduction to the study
of Protozoa. Clarendon Press, Oxford.
P u y t o r a c P. d e 1952: Maupasella glossoscolecis
nov. sp. et M. lubeti nov. sp.
ciliés astomes commensaux du tube digestif de Glossoscolex
grandis
(Michlsm). Remarques sur les Maupasellinae.
Bull. Soc. Zool. Fr., 76, 323.
P u y t o r a c P. d e 1954: Contribution à l'étude cytologique et taxonomique des
Infusoires astomes. Ann. Sci. Nat. Zool. (Ser. 11), 16, 85-270.
P u y t o r a c P. d e 1972: Les ciliés astomes Haplitophryidae
II. Revision de la
systématique dece group. Protistologica, 8, 5-42.
W i c h t e r m a n R. 1939: Investigations on three new ciliates from the littoral
earthworm of Tortugas. Yearb. Carnegie. Inst. Washington, 38
EXPLANATION OF PLATE I
Maupasella perionychis sp. n.
3: Showing arrangement of kineties in ventral view. (Based on silver impregnated preparations
4: Entire organism
5: The entire end magnified to show the triangular naked area
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PLATE I
ACTA P R O T O Z O O L . VOL. 1U, N o . 1
P. A. Lalpotu
auctor phot.
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ACTA
Vol. 19 (No. 1)
PROTOZOOLOGICA
WARSZAWA, 30 III 1980
pp. 61-66
Department of Zoology, Andhra University, Waltair, India
C. C. N A R A S I M H A M U R T I
and S. N a z e e r A H A M E D 2
A New Septate Gregarine, Actinocephalus bradinopygi sp. n.
from the Dragon Fly, Bradinopyga geminata Rambur 1
Received
18 July 1979
Synopsis. The morphology and life-history of a new species of septate
gregarine, Actinocephalus
bradinopygi,
sp. n. from the gut of the odonate, Bradinopyga geminata Ramb. is described.
The cephalonts reach 825 X 168 M,m and the epimerite is complex.
Cysts spherical, 800 pim in diameter and have an ectocyst 150 |.im
thick. Gametes isogamous, 10 nm in diameter. Spores oval, 17X5 pim,
octozoic. Sporozoites spindle-shaped, 10X3fxm.
While examining the local odonate fauna for gregarine parasites we
frequently came across a septate gregarine belonging to the genus
Actinocephalus parasitic in the gut of the dragon fly, Bradinopyga geminata Rambur. A perusal of the literature showed that there has
so far been only one species, A. sieboldi (Kolliker, 1848) reported from
the gut of the larvae and imago of Aeschna sp. and Sympetrum
vulgatum, S. pedemontum
collected at Wurtzburg, Erlangen, Beyrouth.
Gdańsk and Silesia (Poland) (G e u s 1969). The present form differs
from it in several respects and hence is considered a new species for
which the name Actinocephalus bradinopygi sp. n. is proposed.
Material
and
Methods
The dragon flies were collected from the dairy farm area in Visakhapatnam
(Andhra Pradesh, India) where they are found actively flying around a stream
of water.. They were also collected from the Botany gardens in the Andhra
1
Part of the Thesis approved for the award of the Doctor of Philosophy
degree of the Andhra University, Waltair.
2
Present address: Government H. S. School, Pasighat, Arunachal Pradesh,
India.
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62
University Campus at Waltair. The insects were trapped using a net and maintained singly in glass finger-bowls at the bottom of each of which a moist blotting paper was placed to keep the faecal matter passed out in a moist condition.
There was no external indication of infection and hence all the insects collected
were decapitated and the gut examined microscopically for the gregarines. Smears
were prepared from the gut of the infected hosts, fixed in Schaudinn's fluid
and stained with Ehrlich's acid haematoxylin. Cysts collected from the faecal
matter were kept in 2.5% aqueous Potassium dichromate and kept in a moist
chamber to observe gametogenesis and sporogony. Smears of the cysts showing
gametes were fixed in Schaudinn's fluid and stained with Ehrlich's acid haematoxylin while those showing spores were fixed in Carnoy's fluid ana treated
according to Feulgen's technique. Material for sectioning was fixed in alcoholic
Bouin's fluid, sectioned at 8 nm thickness and stained with Heidenhain's iron
haematoxylin.
Observations
38 out of 60 adult Bradinopyga geminata Rambur were infected with
a septate gregarine belonging to the genus Actinocephalus.
T r o p h o z o i t e s : No intracellular stage of development was observed. The smallest cephalont measured 316 X 44 |im and the largest measured 825 X 168 ^im. The body of the cephalont is divisible into three
parts, an anterior complex epimerite which is broad at the base and is
drawn into a long tubular neck. The tip of the neck is expanded into
a shallow cup, the margin of which is produced into 9-11 petaloid lobes
(Fig. 1, 3). When the epimerite was contracted, which was often the case
in smear preparations, it appeared in the form of a bulb-like structure
at the tip of the epimerite (Fig. 1, 2). In fully grown cephalonts the
epimerite measured 272 jim in length and is broadest at the base where
it measured 60 [xm. The neck which was tubular was 16 jxm in width.
The protomerite was hemispherical and measured 90X116 |im. The deutomerite was the longest part of the body and measured 540X125 |im
and has conspicuous striations, which are more prominant along the
sides (Fig. 1, 5). The protoplasm in the deutomerite was more coarsely
alveolated than that in the protomerite which was finely alveolated.
The nucleus has a variable position in the deutomerite and may be situated either in the anterior, middle or posterior end.
Cephalonts attached to the gut epithelium showed that the lobes
of the cup-shaped part of the epimerite are anchored in the epithelial
cells of the midgut (Fig. 1 J). Some of the cephalonts were also seen
moving about freely in the lumen of the gut, but it has not been possible to ascertain if the cephalonts can detach and reattach themselves
to the gut epithelium. The largest sporadin observed measured 1300X
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N E W G R E G A R I N E ACTINOCEPHALUS
BRADINOPYGI
63
Fig. 1. Actinocephalus
bradinopygi
sp. n. 1 — Attachment, 2 — A cephalont —
note the contracted bulb-like part of epimerite, 3 — An enlarged view of the
epimerite, 4 — A sporont, 5 — A part of the deutomerite showing transverse
striations, 6 — Cyst showing 4 nuclei — 2 in each of the two gametocytes, 7 —<
A multinucleate cyst showing peripheral arrangment of nuclei in the gametocytes, 8 — 2 nucleate sporoblast, 9 — 4 nucleate sporoblast, 10 — A sporocyst showing sporozoites
255 (im and was solitary. The deutomerite shows transverse striations
which were more conspicuous than in the cephalonts.
C y s t s : Cysts were spherical having a diameter of 800 jxm and were
opaque white (Fig. 1 6, 7). A gelatinous ectocyst 150 ^tm thick was
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64
present. Cysts passed out along with the faecal matter showed 20-25
nuclei in each of the gametocytes. When the gamete formation was
completed, which was approximately 48 h after the cysts were passed
out, the cysts appeared transluscent.
G a m e t e s : Gametes are isogamous having a diameter of 10 jim.
No locomotor organelle was observed. The cytoplasm was hyaline and
the nucleus which was deeply stained was placed in the centre.
S p o r o g o n y : Sporogony was completed in about 72 h after the
cysts were passed out and the cysts appeared bluish black and float on
the surface. Dehiscence was by simple rupture. Spores were oval with a rigid wall which was impervious to most chemicals and stains. They were oval
measuring 17.0 X 5.0 urn (Fig. 1 8, 9). When the sporozoites were formed they were arranged in two groups of 4 each on either side, with
a small quantity of residual protoplasm in the centre (Fig. 1 10). The
sporozoites are spindle-shaped measuring 10.0X3.0 (im and contain hyaline cytoplasm. A deeply stained nucleus surrounded by a clear halo
was present in the centre.
Discussion
The form described in the present paper has an epimerite which
generally resembles that of Menospora in having a long slender neck
expanded into a large cup at the apex but lacks hooks. It also resembles
the genus Actinocephalus in posessing 9-11 petaloid lobes (instead of
8-10 bifurcate digitiform processes at the apex of the epimerite as
in Actinocephalus)
placed on a long neck unlike in
Actinocephalus
where only a short neck is present. The spores in the present case
are cylindrical and resemble those of Actinocephalus more closely where
they are described as cylindroconical rather than of Menospora where
they are crescentic. In the present work greater importance is attached to the presence or absence of spines in the epimerite, rather than
the length of the neck of the epimerite and as such, the present form
is placed in the genus Actinocephalus. A. sieboldi is the only species
reported so far from odonate hosts (intestine of the larvae of Aeschna
sp.). A. sieboldi reaches a size of 162-188X45 pirn and the epimerite is
composed of 4-6 sharp recurved hooks while in the present form the
fully grown cephalonts reach 825X165 |xm and the epimerite is different. The spores in A. sieboldi are described as biconical and the size is
not given while in the present form they are cylindrical and measure
17.0X5.0 jim. In view of the above mentioned reasons the form descri-
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N E W G R E G A R I N E ACTINOCEPHALUS
BRAD1NOPYGI
bed in the present paper is considered new and the name
bradinopygi sp. n. after the host is proposed.
65
Actinocephalus
ACKNOWLEDGEMENTS
Thanks are due to Prof. K. Hannumantha Rao, Head of the Department of
Zoology for the facilities provided. One of us (SNA) is thankful to the Council
of Scientific and Industrial Research for the award of a Junior Research Fellowship.
We are thankful to Dr. Stanisław L. Kazubski for critically going through
the manuscript and offering many helpful suggestions.
RÉSUMÉ
La morphologie et le cycle d'évolution sont décrites d'une espèce nouvelle
de segment grégarine, Actinocephalus
bradinopygi sp. n. de l'odonate,
Bradinopyga geminata, Ramb.
Les cephalontes sont 825 X 168 ^im et l'épimerite est complexe. Les kystes sont
sphériques, 800 jim dans diamètre, l'épaisseur du externe kyste mesure 150 ftm,
Gamètes similaire, 10 nm dans diamètre spores ovale, 17 X 5 |am. Octozoic. Sporozoite fusiforme, 10X3 [ira.
REFERENCE
G e u s A. 1969: Sporentierchen, Sporozoa, Die Gregarinida der Land- und Süsswasserbewohnenden Arthropoden Mitteleuropas. In: Die Tierwelt Deutschlands
(ed. F. D a h 1), Gustav Fischer Verlag, Jena, pp. 608.
5 — Acta Protozool. 1/80
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ACTA
Vol. 19 (No. 1)
PROTOZOOLOGICA
WARSZAWA, 30 III 1980
pp. 67-76
Department of Cell Biology, Nencki Institute of Experimental Biology, 02-093 Warszawa,
Pasteura 3, Poland
Zbigniew
BARANOWSKI
Kinetics of the Regeneration of Rhythmic Contraction Activity
in Physarum polycephalum
Drops
Received
31 July 1979
Synopsis. The endoplasmic drops isolated from plasmodial veins of Physarum polycephalum,
under appropriate culture conditions, regenerate
rhythmic contraction activity characteristic of the intact Plasmodium
The time indispensable for resumption of contraction activity follows
a normal distribution, while the kinetics of a trigger mechanism for
this activity is an example of elementary process of Poisson's kinetics.
The temperature dependence of the kinetics of a trigger mechanism shows
a phase transition in the range of 16.6-18.4°C. Approximately, the same
temperature transitions were observed for response of Plasmodium to
chemical stimuli and rate of pseudopodia extension (U e d a and K o b a t a k e 1978). Since these results were interpreted as the existence of
two distinct stages in Plasmodium membrane activities, it is reasonable
to suspect that plasmalemma of Physarum polycephalum
is involved
in a trigger mechanism.
Plasmodium of the slime mould Physarum polycephalum is a favourable model for the study of different aspects of motile activity ( K a m i y a 1959, K o m n i c k et al. 1973). Among them, investigations of
different oscillatory phenomena in its contraction behaviour, the nature
of oscillations and their control are of special interest ( W o h l f a r t h B o t t e r m a n n , in press).
Endoplasmic drops of Physarum polycephalum represent a unique
1
Study undertaken under the Research Project MR II.l of Polish Academy
of Sciences. Experiments performed in the Institute of Cytology, University of
Bonn, were supported by the grant of Deutscher Akademischer Austauschdienst
(DAAD).
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68
Z. B A R A N O W S K I
model for investigation of de novo formation of contractile apparatus
and regeneration of its function { I s e n b e r g and W o h l f a r t h - B o t t e r m a n n 1976). Moreover, the model can be easily obtained by puncturing the plasmodial veins. Endoplasm which fills out the veins is
under hyperpressure and therefore it protrudes and forms a drop
after puncture. The drops isolated from the veins, under appropriate
culture conditions undergo a sequence of spontaneous, time-dependent
differentiation processes which finally lead to a new migrating Plasmodium. The sequence consists of regeneration of plasmalemma (W o h 1f a r t h - B o t t e r m a n n and S t o c k e m 1970), endoplasm-ectoplasm
transition which is accompanied with actin polymerization ( I s e n b e r g
and W o h l f a r t h - B o t t e r m a n n 1976), differentiation of actomyosin fibrils and regeneration of plasmalemma invaginations ( G ö t z v o n
O l e n h u s e n e t a l 1979), de novo generation of contraction rhythmicity ( G ö t z v o n O l e n h u s e n and W o h l f a r t h - B o t t e r m a n n 1979), and finally locomotion of Plasmodium. Up to now, the
problem of de novo generation of vitality of the endoplasmic drops was
mainly restricted to the study of morphogenetic processes. The aim
of the present study was to determine the kinetics of de novo generation of oscillating contraction activity of untreated drops and to postulate the localization of a trigger mechanism for the contractile activities in Plasmodium.
Material
and
Methods
The slime mould Physarum polycephalum was cultured as described by C a m p
(1936). The endoplasmic drops (1-1.5 mm in diameter) were obtained by puncturing the veins of frontal region of freely migrating Plasmodium with a steel
needle.
Immediately after their generation from flowing endoplasm the drops were
separated from Plasmodium and placed in a moist chamber on plexiglass surface.
Time dependent events in the contractile behaviour of endoplasmic drops were
controlled tensiometrically using the "contact method" ( W o h l f a r t h - B o t t e r m a n n 1975) under isometric conditions of measurements. The drops collected
during 8-10 h from the same Plasmodium were counted as a single sample.
The number of drops in the samples varied from 23 to 32.
The duration of Plasmodium migration without feeding before the production
of drops was controlled only in the experiments in which the influence of starvation on de novo generation of rhythmic contraction activity of drops was investigated. The measurements were taken in room temperature (about 21°C).
Effect of temperature on contraction behaviour of the drops was investigated
in a chamber of which temperature was controlled with the aid a Peltier element.
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CONTRACTION ACTIVITY OF
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69
R e s u l t s
A typical tensiometer curve of the contraction behaviour of an endoplasmic drop is shown in Fig. 1. First minimum of visible oscillations
was chosen as the start point of rhythmic contraction activity (stage
S3) of a drop. Duration of preliminary stage is marked as S12.
Fig. 1. A typical tensometric curve of de novo generation of rhythmic contraction activity in an endoplasmic drop of Physarum
polycephalum.
Time of the
drop generation is the starting poin of the measurement. Phases S 12 and S 3
denote preliminary and rhythmic contraction activity stages respectively. S t and
S, — subphases of the preliminary stage
Fig. 2. The histograms of the kinetics of (a) Sir-^Sa (b) — Si->S 2 and (c) —
Sî->S, transitions. N 0 — total number of the drops; N12, N, and N 2 — number
of the drops in S12, Sj and S 2 stages respectively. In the case of S 2 -»S, transition the drops in the sample were synchronized in respect to the begining of
S 2 stage
Figure 2a presents an example of histogram of S12 stage duration,
it means, of kinetics of the transition from preliminary stage (S12) to
oscillation pattern (S3) of contraction activity. The sample consisted
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70
Z. B A R A N O W S K I
of 31 drops and was measured in room temperature. The histogram
is satisfactory approximated by S-shape curve of distribution function
of normal distribution. The approximation curve was obtained on the
basis of the best fit straight line on normal distribution control chart.
Normal distribution of the initiation of rhythmic contraction activity
suggests that the start of oscillations against the time of the drops
generation is a result of numerous independent events which additively
lead to active stage of drops ( M a r t i n 1967).
Tensiometric curves enable to find more elementary stages in contraction behaviour of the drops. In preliminary stage, the drops show
characteristic maximum of the force exerted on the measuring device
receptor. This allows for the arbitral division of Si2 stage into two separate stages Sj and S2, as it was marked in Fig. 1.
Histogram of the time relation of Si
S2 transition (Fig. 2 b) still
suggests a complex character of the transition, that means, is well approximated by S-shape curve.
In order to estimate the duration of S 2 stage and the time relation
of S 2 -» S 3 transition the time point of the maximum of the force exerted on the measuring device receptor was chosen as a time zero for this
distribution (Fig. 2 c). The attempt of approximation of this distribution
by S-shape curve did not give a satisfactory result. With the purpose
of attaining a good approximation the distribution was tested in In N^Nq
and time coordinates, and the best fit straight line was found. Simple
transformation of an equation of a straight line in semilogharitmic coordinates allows to obtain an equation of exponential curve, which in
good manner approximates experimental distribution. Exponential distribution is an example of transitions between stages of Markov processes chain ( B h a r u c h a - R e i d 1960). Simple negative exponential
one is an example of transition between two initial stages (Po and Pi)
of Poisson's kinetics. Theoretically transition P 0 -> Pi is given by equation
P 0 /Pi = exp (— Xt). The transition S 2
S 3 is approximated by N S 2 /N 0 =
= exp
A(t—0)] curve. Thus, the parameter 0 can be interpreted as
a delay in S 2 stage (B 1 i o k h and S m o l y a n i n o v 1977).
The above considerations point out, that the start of oscillation pattern of contraction activity depends on numerous events in initial period
(Si) of the drops behaviour, while terminal one (S2) is determined by
trigger system, which kinetics can be described as a single Poisson's
process.
Investigations of stochastic processes raise the question in what extent a sample can be regarded as representative for population. In other
words, what conditions have to be fulfilled during collecting drops which
make a sample. In order to give some impression concerning this pro-
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C O N T R A C T I O N A C T I V I T Y OF
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71
Fig. 3. The influence of the period of Plasmodium starvation on the kinetics
of the transitions between elementary stages in contraction behaviour of Physarum polycephalum
drops. The curves approximate the histograms of Sj 2 ->S 8 ,
and Sg—>-Ss transitions for samples taken after 12 h (N 0 — 24 drops —
continuous line), 36 h (N 0 = 23 drops, dot-dash line) and 60 h (N 0 = 32 drops —
dotted line) of starvation. N12, Nx and N2 — number of the drops in S 12 ,
S! and S 2 stages respectively
blem the kinetics of starting point of rhythmic contraction activity was
investigated at different periods of Plasmodium starvation. The drops
were collected after 12, 36 and 60 h of starvation of the same Plasmodium. The results are shown in Fig. 3. There are drawn the best fit
curves for S12 -> S3, Si -> S 2 and S 2
S 3 transitions. The curves were
obtained in the same manner as in the case of distributions shown in
Fig. 2.
Since Sia
S 3 and Si -> S 2 transitions follow a normal distribution,
their means equality can be tested by Student's difference test ( B r a n d t
1970). The hypothesis of equal means has to be rejected only in the case
of Si2 -> S 3 transition for samples taken after 12 and 60 h of starvation
(!t| = 2.88 > P0.05 = 2.01). This result points that an intrinsic process
or processes taking place in Plasmodium during starvation can change
parameters of distributions. It mainly concerns the terminal phase of
S12 S 3 transition, thus, a postulated trigger system, which acts during
S2
S 3 transition. On the other hand it imposes conditions of sample
making for investigation of the kinetics of a trigger system. Drops have
to be collected from a single Plasmodium no longer than during 12 h,
because otherwise slight differences in parameters of negative exponential distribution would lead to S-shape of distribution function of
S 2 ->S 3 transition.
The kind of kinetics of above mentioned transitions does not change
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72
Z. B A R A N O W S K I
Table 1
Temperature dependence of mean time of transitions between elementary stages in contraction
behaviour of Physarum polycephalum drops
T
[°q
15.8
16.6
18.4
19.3
20.7
23.7
Sl2 - > S3
f+Sx min
49.5±4.5
44.3±2.5
21.7±0.9
19.3±0.6
15.6±0.3
10.2±0.4
St->S 2
S
10.1
5.6
3.9
4.3
2.6
2.4
f+Sx min
15.4±2.3
11.8±1.3
7.7±0.4
7.6±0.4
7.6±0.4
4.2±0.2
s2->s3
s
5.2
2.9
2.0
2.6
2.0
1.0
f+Sx
min
34.3±3.3
32.5+3.1
14.3±0.8
11.9+0.5
7.9+0.6
5.9+0.4
S
7.4
7.1
3.8
3.5
3.2
2.1
S — standard deviation of the samples, S x — standard error of the samples
Fig. 4. T e m p e r a t u r e d e p e n d e n c e of m e a n t i m e of Si2->S 8 , S ^ S a a n d S 2 -vS 3 t r a n sitions (see Table 1) plotted in s e m i l o g h a r i t h m i c c o o r d i n a t e s
with temperature, that means, they preserve their S-shape or, respectively, negative exponential character. It was tested at 18.4, 19.3, 20.7,
and 23.7°C. However, there exist conspicuous differences between mean
time and standard deviations of all transitions (Table 1).
Figure 4 shows the same data plotted as In tsi2, s1? s2 against temperature. There is a clear discontinuous change of the mean time values of
Sia -> S3 and S 2 —»• S 3 transitions, while there is no such in Si -> S2 transition. The mean time of initiation of rhythmic contraction activity
against the time of drops generation (tsl2), and the time of intervention
of a trigger mechanism (ts2), are decreased by factor 1/2 in the range of
16.6-18.4°C.
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C O N T R A C T I O N A C T I V I T Y OF
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73
Discussion
The presented results concerning kinetics of de novo generation of
rhythmic contraction activity of the endoplasmic drops (Si2
S 3 transition) confirm a complexity of processes which finally lead to contraction behaviour characteristic of the intact Plasmodium. The complexity
is reflected by normal distribution of this transition. In preliminary stage
the drops undergo very quick process of plasmalemma regeneration by
means of membrane vesiculation ( W o h l f a r t h - B o t t e r m a n n and
S t o c k e m 1970). The organization of the contractile gel reticulum of
a drop and regeneration of plasmalemma invaginations is over after
about 5-10 min ( I s e n b e r g and
Wohlfarth-Bottermann
1976, G ö t z v o n O l e n h u s e n
e t a l . 1979). Then, the drops regenerate rhythmic pattern of contraction activity ( G ö t z v o n O l e n h u s e n and W o h l f a r t h - B o t t e r m a n n 1979).
On the basis of listed above sequence of time-dependent processes
the Sx stage of the drops (Fig. 1) with mean time of 7.6 ± 0.4 min at
room temperature can be interpreted as complex of processes of regeneration of plasmalemma, actin transformations and fibrilogenesis. It
is additionally confirmed by S-shape curve (Fig. 2 and Fig. 3) of Si —v
Sa transition. Simultaneously, S2 seems to be a stage, when contractile
apparatus of the drops, from morphogenetic point of view, is ready for
initiation of contraction activity and is waiting for a stimulus. Thus,
S2
S 3 transition shows the kinetics of a trigger mechanism which stimulates the contractile system of Physarum polycephalum. The kinetics
of a trigger, as it is shown in Fig. 2 and Fig. 3, is represented by
a single Poisson's process. Elementary stages of Poisson's kinetics were
also found in a process of fibroblasts spreading (B 1 i o k h and S m ol y a n i n o v 1977). However, the relationship between actomyosin fibrilogenesis and generation of a force (Fig. 1) at the preliminary stage
of a drop remains obscure at the present time.
The study of kinetics of Si2 -> S 3 and Si
S2 transitions and of
a trigger mechanism against the time of Plasmodium starvation was
done in order to define the homogeneity of the samples. Period of starvation was chosen arbitraly as a factor influencing many inner processes taking place in Plasmodium, which can disturb the kinetics of investigated transitions. The obtained results point out that the kinetics
of a trigger system is sensitive to the period of Plasmodium starvation.
Simultaneously, more or less "pure" kinetics of a trigger system can
be investigated when drops are collected from the single Plasmodium
no longer than during 12 h.
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74
Z. B A R A N O W S K I
The distinct differences between mean time of de novo generation
of rhythmic contraction activity are observed with temperature variations (Table 1 and Fig. 4). In the range of 16-24°C, mean time of Si 2 ->
S3, Si
S 2 and triggering of oscillations decrease exponentialy (Fig. 4)
with the increase of temperature. However, the curves for S J2
S 3 and
a trigger mechanism show discontinuous decreasing by factor 1/2 in
range of 16.6-18.4°C. It means that the trigger mechanism of de novo
generation of oscillation contraction activity has two distinct stages.
A phase transition between these stages is caused by temperature variations.
Discontinuous changes in magnitude of the response of plasmodia to
chemical stimuli and the rate of pseudopodia extension caused by temperature variations were found by U e d a and K o b a t a k e (1978).
The transition temperature for these phenomena is about 15°C. The results were interpreted as a conformational change of the surface membrane of Physarum polycephalum at transition temperature.
Plasmalemma was earlier suspected to be a site of a trigger mechanism which acts via changes in ionic permeabilities and/or electrogenic ion pumps. However, still there is a lack of clear experimental
evidence of this hypothesis (W o h l f a r t h - B o t t e r m a n n in press).
Conformity of transition temperatures for chemotactic phenomena, rate
of pseudopodia extension and time of intervention of a trigger mechanism might suggests that plasmalemma of Physarum, polycephalum is
involved in a trigger system, but the present results give no background to formulate any hypothesis explaining the mechanism of this
system.
ACKNOWLEDGMENTS
My thanks are due to Prof. K. E. Wohlfarth-Bottermann, Head of the Institute of Cytology and Micromorphology, University of Bonn for giving necessary
facilities to carry out the t e n s o m e t r i c measurements and for the helpfull discussions.
RÉSUMÉ
Les gouttes d'endoplasme isolées des veines du Plasmodium de
Physarum
polycephalum,
maintenues dans les conditions ambiantes favorables, récupèrent
l'activité contractile rythmique qui est caractéristique pour un Plasmodium intact.
La variation du temps nécessaire pour la restitution de l'activité contractile suit
une
distribution normale, tandis que la
cinétique
du
mécanisme
qui
déclenche
cette
activité
représente
le
cas
d'un
processus
élémentaire
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C O N T R A C T I O N A C T I V I T Y OF
PHYSARUM
75
obéissant à la cinétique de Poisson. La dépendence de la cinétique du mécanisme
déclencheur de la température manifeste une transition de phase entre 16.6 et
18.4°C. Une transition thermique approximetivement pareille était observée pour
la réponse du plasmodium à la stimulation chimique et la vitesse d'extension
des pseudopodes ( U e d a and K o b a t a k e 1978). Ces résultats sont interprétés
comme dus à la présence de deux stades distinctes de l'activité de la membrane
plasmodiale, on peut donc raisonablement supposer que la plasmalemma de P h y sarum polycephalum
est engagée dans le fonctionnement du mécanisme déclencheur.
REFERENCES
B h a r u c h a - R e i d A. T. 1960: Elements of the theory of Markov processes
and their applications. McGraw-Hill Book Company, Inc., N e w York, Toronto, London.
B l i o k h Z h . L. and S m o l y a n i n o v V. V. 1977: Kinetics of fibroblasts spreading. Biophysica, XXII, 281-288.
B r a n d t S. 1970: Statistical computational methods in data analysis. North-Holland Publishing Company-Amsterdam and London, and American Elsevier
Publishing Company, Inc.-New York.
C a m p W. G. 1936: A method of cultivating myxomycete Plasmodia. Bull. Torrey
Bot. Club, 63, 205—210.
G ö t z v o n O l e n h u s e n K., J ü c k e r H. and W o h l f a r t h - B o t t e r m a n n
K. E. 1979: Induction of a plasmodial stage of Physarum without plasmalemma invaginations. Cell. Tiss. Res., 197, 463-477.
G ö t z v o n O l e n h u s e n K. and W o h l f a r t h - B o t t e r m a n n K. E. 1979:
Effects of caffeine and I^O on persistence and de novo generation of intrinsic oscillatory contraction automaticity in Physarum.
Cell Tiss. Res.,
197, 479—499.
I s e n b e r g G. and W o h l f a r t h - B o t t e r m a n n K. E. 1976: Transformation
of cytoplasmic actin. Importance for the organization of the contractile gel
reticulum and the contraction-relaxation cycle of cytoplasmic aotomyosin.
Cell Tiss. Res., 173, 495-528.
K a m i y a N. 1959: Protoplasmic streaming. Protoplasmatologia, VIII, 3, Wien:
Springer.
K o m n i c k H., S t o c k e m W. and W o h l f a r t h - B o t t e r m a n n K. E. 1973:
Cell motility: mechanisms of protoplasmic streaming and ameboid movement. Int. Rev. Cytol., 34, 169-249.
M a r t i n J. 1967: Nations de base en mathematiques et statistiques. Gauthier-Villars, Paris.
U e d a T. and K o b a t a k e Y. 1978: Discontinuous change in membrane activities
of Plasmodium of Physarum polycephalum
caused by temperature variation:
effects on chemoreception and ameboid motility. Cell Struct. Funct. 3,
129-139.
W o h l f a r t h - B o t t e r m a n n K. E. and S t o c k e m W. 1970: Die Regeneration
des Plasmalemmas von Physarum polycephalum.
Wilhelm Roux'Archiv., 164,
321-340.
W o h l f a r t h - B o t t e r m a n n K. E. 1975: Tensometric demonstration of endogenous oscillating contractions in Plasmodia of Physarum
polycephalum.
Z. Pflanzenphysiol., 76, 14-27.
W o h l f a r t h - B o t t e r m a n n K. E. Oscillating contraction activity in Physarum. Workshop on cellular oscillators, Titisee 1979. J. Exp. Biol, (in press).
http://rcin.org.pl
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ACTA
Vol. 19 (No. 1)
PROTOZOOLOGICA
WARSZAWA, 30 III 1980
pp. 77-82
Department of Cell Biology, Nencki Institute of Experimental Biology, Pasteura 3 , 02-093
Warszawa, Poland
Barbara
TOLLOCZKO
Cytochalasin B Induced Inhibition of Food Ingestion
in Dileptus anser (Ciliata, Gymnostomata)
Received
7 August 1979
Synopsis. The influence of different concentrations of cytochalasin B
on Dileptus
anser was studied. The physiological observations
ultrastructural studies were carried out. The inhibitory effect of
drug on food vacuole formation process was stated. The possible
chanism of CB action was discussed.
(CB)
and
this
me-
The cytochalasins are a group of fungal metabolites which have
been found to have inhibitory effects on a variety of cell processes.
The effects of CB are primarily of two types: (1). It has been proposed
that it causes the disintegration of microfilaments and myofilaments
< S c h r o e d e r 1970, B l u e m i n k 1971, S p o o n e r et al. 1971, W e s s e l s et al. 1971, S p o o n e r 1973, A x l i n e and R e a v e n 1974, C op e l a n d 1974, S a c h s et al. 1974, W i l l i a m s o n 1975). Some biochemical studies have postulated a direct interaction of cytochalasins
with actin and/or myosin ( S p u d i c h 1972, S p u d i c h and L i n 1972,
H a r t w i g and S t o s s e 1 1976). (2). Other authors, however, have not
accepted these conclusions and have proposed alternative explanation
such as membranotropic effects for the mechanism of cytochalasins
action ( H a u s c h k a 1973, R a t h k e et al. 1975, W e b e r et al. 1976).
Cytochalasin B (which has beeen used in most studies) has been
found to inhibit phagocytosis in various cells (W i 11 i a m s and W o l f
1971, Z i g m o n d and H i r s c h 1972, A x l i n e and R e a v e n 1974,
N i 1 s s o n et al. 1973, H o f f m a n et al. 1974, R o t h s t e i n and B l u m
1974, N i l s s o n 1977). Since the food ingestion in carnivorous Ciliate — Dileptus anser involves both: membrane and filament activity
and each of these structures is mainly involved in different stage of
the process, the observations of CB influence on different stages of
endocytosis can help in the elucidating of the way of CB action. On
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78
B.
TOLtOCZKO
the other hand the observations of physiology and ultrastructure of
the cell after CB treatment may explain the mechanisms involved in
the process of food ingestion in Dileptus.
Material
and
Methods
Dileptus anser O. F. M. was used in this study. The cells were cultured
in Pringsheim solution and fed every other day with Colpidium colpoda. For
experiments the cells before feeding were selected. They were washed in 2 M M
phosphate buffer pH 7.1 overnight. Stock solution of 10 mg of CB (Serva) in 1 ml
of DMSO was prepared. The CB solutions used in experiments were prepared by
using ImM phosphate buffer pH 7.1. The experiments were carried out at room
temperature. 5-20 fig/ml CB was added to the samples containing about 20 cells
each. The appropriate solutions of DMSO (0.5-2°/o) and phosphate buffer were
used as control media. After 5, 30, 60 min and 24 h of CB, DMSO, or buffer
treatment Colpidia were added in order to test the food ingestion ability of
Dileptus cells. The number of cells containing food vacuoles and cells without
vacuoles after 5 min or 30 min exposure to the prey was counted. The experiments
have been repeated ten times and the mean number (in per cent) of the cells
with, and without vacuoles was calculated. The observations of the single, living
cells treated with CB and exposed to the prey were also carried out.
In order to test the influence of CB on the ultrastructure of Dileptus the
preparations of CB treated and control cells for electron microscopy were made.
The fixation method of G o l i A s k a (1978) was used. The sections obtained by using LKB ultramicrotome were contrasted with uranyl acetate and lead citrate and
examined in JEM 100 B electron microscope.
R e s u l t s
The treatment of Dileptus anser cells with CB causes the inhibition
of endocytosis. The effect is stronger in higher concentrations of the
drug (Table 1, 2) and it is more distinct when the data obtained during
5 min exposure to the food are taken into account (Table 1). This observation indicates that the complete inhibition of food ingestion does not
occur, but the process is much slower then in control cells. The rate
of endocytosis in DMSO treated cells is little slower than in buffer
medium, however, it is significantly higher than in CB treated cells.
No significant differences are seen in the food ingestion ability depending on different time of CB treatment except of 24 h exposure to the
drug, when the complete inhibition of vacuoles formation occurs.
The observations of the behaviour of living Dileptuses treated with
CB have revealed that only the second step of endocytosis is inhibited
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DILEPTUS
Time of
treatment
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3
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o 00
1 in (N NO
Time of
treatment
The percentage of cells of Dileptus anser containing food vacuoles (+) and without food vacuoles (—) after different time of CB,
DMSO or phosphate buffer treatment. Colpidia added for 5 min
Table 1
I N H I B I T I O N OF F O O D I N G E S T I O N I N
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by this agent. It means that the predators do not loose their ability to
capture the prey cells, but they can not ingest them.
The studying of the ultrastructure does not reveal very distinct differences between the microfibrillar ring which holds together small
nemadesmata of the internal basket of oral apparatus in CB treated
and control cells (PI. I 1, 2) and in microfibrillar layer situated between
ecto-and endoplasm in the whole body of Dileptus.
Discussion
The effects of CB on food ingestion in Dileptus a user can be due
to its action either on the membrane or on the microfilaments involved in food vacuole formation. The observations of the living cells
after CB treatment have revealed that the process of opening of the
mouth and vacuole formation is effected by CB. It has been suggested
that in opening of mouth the microfilament ring in which the internal
nemadesmal basket is anchored, can play a role ( T o l l o c z k o 1979).
It has been also postulated by the same author that the new filament
material appears during food ingestion in the area of the oral apparatus.
On the other hand a lot of membrane material in form of smooth,,
elongated vesicles has been observed in endoplasm inside the nemadesmal basket of Dileptus anser (G o 1 i n s k a 1978). These vesicles are
probably the pool of the membrane which is used during vacuole formation similar as in Paramecium ( A l l e n 1974). Since no vacuole formation was observed in CB treated Dileptuses the fusion of the vesicles
with the cytostomal membrane does not occur after CB treatment. On
the contrary, in noncarnivorous Ciliates Paramecium and Tetrahymena
enormously big vacuoles were formed in CB treated cells, but they
were not able to separate (N i 1 s s o n 1973, N i 1 s s o n et al. 1973, T o 11 o c z k o 1977). These observations suggest that some CB sensitive mechanism acts in Dileptus prior to the vacuole formation, whereas in
Paramecium and Tetrahymena it appears during food vacuole separation. It can be assumed that it could be microfilament action. This
assumption is supported by many observations that CB acts on many
filamentous structures (see Introduction). On the other hand, the observation that CB acts on food ingestion already after 5 min treatment is
in agreement with studies of H a u s c h k a (1973) who has postulated
partitioning of CB in hydrophobic regions of the cell membrane. Since
ultrastructural differences between CB treated and untreated Dileptuses are not clearly seen — the most possible explanation is that the effects
of CB are due to its influence first on the membrane and then on
microfilaments action, but without causing distinct differences in their
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INHIBITION OF FOOD INGESTION IN
DILEPTUS
81
ultrastructure. Similar data have been obtained by B o h a t i e r and
K i n k (1977) who observed unnormal function of regenerated, normally
built oral apparatus, when cyclohexymide was applicated during regeneration of Dileptus anser. R a t h k e et al. (1975) have observed no microfilament disintegration during primary phase of CB action. K u r k i n e n et al. (1978) have also observed the influence of this drug on
both: membrane phenomena and actin containing microfilaments in
cultured fibroblasts.
The hypothesis of W e b e r et al. (1976) that cytochalasins induce
the changes in plasma membrane and these changes cause as a second
step the changes in microfilaments seems to be the most probable explanation of CB action on food ingestion process in Dileptus.
ZUSAMMENFASSUNG
Der Einfluss
tus anser wurde
den durchgefürt.
handlung wurde
chprochen.
von diversen Konzentrationen von Cytochalasin B (CB) auf Dilepunterschut. Physiologische und ultrastrukturalen Forschungen wurDie Hemmung von Ernahrungsvakuolenbildung nach der CB Befestgestellt. Möglicher Mechanismus von CB-aktion wurde bes-
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W i l l i a m s o n R. E. 1975: Cytoplasmic streaming in Chara: A cell model activated by ATP and inhibited by cytochalasin B. J. Cell. Sci., 17, 655-368.
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73, 383-393.
EXPLANATION OF PLATE I
Fragments of microfibrillar ring in which nemadesmal basket is anchored, n —
nemadesma, m — microfilaments
1: Control cell of Dileptus anser (X54 000)
2: Dileptus anser after 1 h treatment with 10 fxg/ml CB (X54 000)
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A C T A PROTOZOOL. VOL. 19, Mo. 1
PLATE I
auctor phot.
B. Tolloczko
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In
preparation:
S. F a b c z a k : Electrical Properties of Cell M e m b r a n e in Protozoan,
Stentor
coeruleus.
I. Modification of Resting M e m b r a n e Potential by Extracellular
Ions — S. F a b c z a k : Electrical Properties of Cell Membrane in Protozoan
Stentor coeruleus. II. Effect of T e m p e r a t u r e on I n t e r n a l M e m b r a n e Potential —
A. W a s i k and J. S i k o r a : Effects of Cytochalasin В and Colchicine on Cytobursaria — B. T o ł ł o c z k o : Effect of Colplasmic Streaming in Paramecium
chicine on Food Ingestion in Dileptus anser — Z. S a y e r s : Motile Behaviour of
Amoeba proteus under Various Ionic Conditions — W. K ł o p o c k a and A. G r ę b e c k i : Motory Independence of Pseudopodia in Freely Moving Amoeba proteus —
M. C i e ś l a w s k a : Dynamics of the Ending Veins in Plasmodia of
Physarum
polycephalum
— J. K o ł o d z i e j c z y k and A. G r ę b e c k i : Dynamics of the
F r o n t a l Margin of Plasmodia of Physarum
polycephalum
— B. S k o c z y l a s
and D. K u c h a r c z y k : Reproducibility of Paramecium
tetraurelia
strain 51 S
and Paramecium
octawrelia strain 299 S in Mass Cultures — V. G o l e m a n s k y
and D. K u l d j i e v a : Eimeria garzettae sp. n. (Coccidia: Eimeriidae) in the Little
Egret (Egretta garzetta L.) f r o m Bulgaria — M. M u k h e r j e e and D. P. H a 1d a r : Observations on Eimeria glossogobii sp. n. (Sporozoa: Eimeriidae)
from a
Fresh Water Teleost Fish — А. К о в а л ь ч у к : Быстрый метод изготовления постоянных препаратов инфузорий в экспедиционных условиях.
Warunki p r e n u m e r a t y
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Cena zł 50.—
CONTENTS
G.
M.
M.
W.
A.
P.
C.
Z.
B.
A. A n i t i p a : A Temporal Analysis of Cell Cycle and Morphogenese
Events in T etrahymena
pyriformis
" W o l s k a : Tetratoxum
unifasciculatum
(Fiorent.) (Ciliata,
Entodiniomorphida). I. Somatic and Adoral I n f r a c i l i a t u r e
W o l s k a : Tetratoxum
unifasciculatum
(Fiorent.) (Ciliata,
Entodiniomorphida).
II. Electron Microscope Investigations
F o i s s n e r : Taxonomische Studien über die Ciliaten des Grossglocknergebietes (Hohe Tauern, Österreich). VI. Familien Woodruffiidae,
Colpodidae und
Marynidae
[Taxoncmical Studies on the Ciliates of t h e Grossglockner Area (Hohe
Tauern, Austria). VI. Families Woodruffiidae,
Colpodidae,
and
MarynidaeJ
K. C h a n d r a and A. C h o u d h u r y : A New Opalinid Hegneriella
mukundai sp. n. f r o m an Old World Hylid Kaloula pulchra
taprobanica
P a r k e r from India
A. L a l p o t u : Maupasella perionychis
sp. n. a New Astomatous Ciliata
from the E a r t h w o r m Perionyx
sensibaricus
in India
C. N a r a s i m h a m u r t i and S. N a z e e r A h a m e d : A New Septate
Gregarine Actinocephalus
bradinopygi
sp. n. f r o m the Dragon Fly, Bradinopyga geminata R a m b u r
B a r a n o w s k i : Kinetics of t h e Regeneration of Rhythmic Contraction
Activity in Physarum polycephalum
Drops
T o l l o c z k o : Cytochalasin B Induced Inhibition of Food Ingestion in
Dileptus anser (Ciliata, Gymnostomata)
1
15
21
29
51
55
61
67
77
Państwowe Wydawnictwo Naukowe — Oddział we Wrocławiu
Wrocławska D r u k a r n i a Naukowa
Index 35133
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